Adenylosuccinate lyase (ADSL) is a conserved homotetrameric enzyme that catalyzes the eighth reaction in the de novo purine synthesis (DNPS) pathway and also participates in the purine nucleotide cycle (Figure 1—figure supplement 1; Daignan-Fornier and Pinson, 2019). Mutations in ADSL cause adenylosuccinate lyase deficiency (ADSLD), an autosomal recessive disorder characterized by defects in purine metabolism and heterogeneous neurological phenotypes that include lack of eye-to-eye contact, auto-aggressive behavior, speech impairment, mild psychomotor delay, transient contact defects, autism spectrum disorder, epilepsy, and in some cases, microcephaly, encephalopathy, ataxia, or coma vigil (Jaeken and Van den Berghe, 1984; Jurecka et al., 2015). While the incidence of ADSLD has not been fully established, over 100 patients have been diagnosed to date and subcategorized based on their symptoms that range from premature death to milder developmental and behavioral disorders (Jurecka et al., 2015; http://www.adenylosuccinatelyasedeficiency.com/).

ADSLD can be diagnosed by detecting elevated levels of the substrates of ADSL, SAICAR (also referred to as SZMP), and S-AMP, as well as their dephosphorylated riboside forms, SAICAr and S-Ado, in bodily fluids (Jaeken and Van den Berghe, 1984). As normal levels of purine nucleotides were detected in serum from ADSLD patients, the accumulation of S-Ado, and particularly SAICAr, was proposed to play a role in the disease pathology (Jaeken and Van den Berghe, 1984; Jurecka et al., 2015; Stone et al., 1998).

In yeast, ADSL (Ade13) loss provokes genomic instability and is lethal (Daignan-Fornier and Pinson, 2019; Giaever et al., 2002; Pinson et al., 2009; Chen et al., 2016). Lethality in yeast can be rescued by deletion of a number of DNPS enzymes upstream of ADSL, or the transcription factors that regulate the pathway, indicating that the accumulation of metabolic intermediates, rather than impaired DNPS, underlies the toxicity (Daignan-Fornier and Pinson, 2019; Chen et al., 2016). In Caenorhabditis elegans, ADSL loss caused delayed growth, infertility, reduced life span, and locomotion defects. In some studies, growth, life span, and locomotion could be linked to the accumulation of SAICAR (Chen et al., 2016; Marsac et al., 2019; Fenton et al., 2017).

Perfusion of rat brains with SAICAr caused cellular attrition in the hippocampus, leading to the proposition that SAICAr accumulation is neurotoxic, although the potential mechanism remains unknown (Stone et al., 1998). In glucose-deprived cancer cells, SAICAR accumulation was shown to activate PKM2, and a number of other kinases, to promote cancer survival in glucose-limiting conditions, suggesting that purine metabolite accumulation could have distinct signaling outcomes that impact on cell behavior and fate during development (Keller et al., 2012; Keller et al., 2014; Yan et al., 2016). However, despite extensive enzymology and structural information, the underlying mechanisms by which neuropathology arises in ADSLD remain unknown.

To address the potential roles of ADSL deficiency in neurodevelopment, we systematically examined the consequences of ADSL depletion in diploid human cells and in vivo. We found that ADSL depletion in human epithelial cells impaired cell cycle progression, induced DNA damage signaling, and impaired primary ciliogenesis. Deletion of p53 or supplementation with nucleosides rescued cell cycle progression and DNA damage signaling, respectively. In contrast, ciliogenesis defects were unaffected by p53 status or nucleoside supplementation and could be reproduced by SAICAr supplementation or attenuated by the inhibition of phosphoribosylaminoimidazole carboxylase (PAICS), which is required to generate SAICAR/r upstream of ADSL. Depletion of ADSL in chicken or zebrafish embryos impaired neurogenesis and caused developmental defects. Both chicken and zebrafish embryos exhibited microcephaly, which is observed in some ADSLD patients, and increased DNA damage was evident in zebrafish. In addition, fish embryos displayed ciliopathy-related phenotypes and interventions to limit SAICAR accumulation-rescued impaired neurogenesis. Together, our results indicate that ADSL depletion causes context-dependent phenotypes associated with its role in both the purine nucleotide cycle and DNPS, which together impact neurodevelopment.

Despite a detailed understanding of the enzymology of DNPS and the purine nucleotide cycle, the specific cell and organismal effects underlying the complex etiology of ADSLD remain unclear. Our results uncovered multiple phenotypes associated with ADSL depletion in human RPE-1 cells, as well as developing chicken and zebrafish embryos (Appendix 1). These can largely be rescued by interventions that suppress the generation of metabolites in the DNPS pathway or restore purine levels, complementing defective DNPS or purine nucleotide pathway activity. DNA damage signaling was suppressed by nucleoside supplementation in human cells and zebrafish, suggesting that this was caused by purine deficiency (Figure 1A). In contrast, defects in primary ciliogenesis resulting from ADSL depletion were rescued by PAICS inhibitor or MTX treatment, but not nucleosides, and phenocopied by SAICAr administration. The inability to rescue these phenotypes with nucleoside supplementation indicated that they most likely resulted specifically from loss of ADSL, and potentially SAICAr accumulation, rather than a general deficiency in purine supply. As SAICAr levels were barely detectable in RPE-1, it could indicate that a very small amount of SAICAr is sufficient to provoke the phenotypes or that signaling elicited specifically from loss of the ADSL protein is responsible. In ADSL-depleted RPE-1 cells, we could also detect p53 activation and defects in cell cycle progression, in the absence of cell death or senescence, and these phenotypes were insensitive to nucleoside supplementation or DNPS modulation, indicating the involvement of additional pathways.

Microcephaly, which is present in a subset of more severe ADSLD cases (Jurecka et al., 2008; Jurecka et al., 2012; Mouchegh et al., 2007), was observed in zebrafish embryos following Adsl depletion. Similarly, ADSL depletion in chicken embryos led to a reduction in neural tube size. Together, the results suggest that they are potentially valuable models for understanding the etiology of microcephaly in ADSLD (Jurecka et al., 2015). DNA damage, p53 activation, and defects in cilia function, which we observed following ADSL depletion, have all been implicated in neurodevelopmental disorders, such as Seckel syndrome, which is associated with microcephaly (Stracker et al., 2020). In Seckel syndrome mice expressing hypomorphic, humanized alleles of Atr, replication stress and DNA damage preceded neuroprogenitor cell death (O’Driscoll et al., 2003; Murga et al., 2009). This was accompanied by extensive p53 activation, but co-deletion of p53 exacerbated the cellular and organismal phenotypes, indicating a protective effect of p53 induction. Mutations in centrosomal proteins, such as CENPJ/SAS4/CPAP or CEP63, were also implicated in Seckel syndrome (Al-Dosari et al., 2010; Sir et al., 2011). In contrast to mice expressing hypomorphic Atr, progenitor loss in mice with CEP63 or CENPJ/SAS4 deficiency resulted from mitotic delays and the activation of the USP28-53BP1-dependent mitotic surveillance pathway (Bazzi and Anderson, 2014; Insolera et al., 2014; Marjanović et al., 2015; McIntyre et al., 2012; Lin et al., 2020; Fong et al., 2016; Lambrus et al., 2016). In this case, the phenotype was completely rescued by p53 co-deletion, revealing p53-dependent cell death as a main driver of the phenotype. Despite some phenotypic similarities with Seckel syndrome at the cellular level, including replication stress and p53 activation, we did not detect increased cell death as a result of ADSL depletion. This indicated that the reduced cellularity caused by ADSL depletion occurs in a manner mechanistically distinct from Seckel syndrome driven by ATR deficiency or centrosome duplication defects.

GMPS (GMP synthase), an essential enzyme in the purine synthesis pathway (Figure 1—figure supplement 1), was identified as a target for p53 repression following p53 activation using Nutlin-3a (Holzer et al., 2017). In addition, GMPS was demonstrated to promote p53 stabilization in response to genotoxic stress in a manner that requires the USP7 ubiquitin protease (Reddy et al., 2014). While we cannot rule out a role for this pathway in p53 stabilization following ADSL depletion, we could not rescue p53 activation with nucleoside supplementation and induction of p53 via the GMPS pathway was demonstrated to induce cell death, which we did not observe clearly in any of the systems we analyzed.

In addition to p53-dependent and -independent cell death, defects in cilium removal and premature progenitor differentiation have also been clearly implicated in microcephaly in Seckel syndrome and primary microcephaly (Gabriel et al., 2016; Farooq et al., 2020). We examined the removal of cilia by adding back serum to serum-starved RPE-1 cells depleted for ADSL and observed normal rates of cilia removal (Figure 4—figure supplement 1G), suggesting that impaired cilia resorption was likely not impairing cell cycle exit and indirectly impacting ciliogenesis. Analysis of differentiation in RPE-1 cells depleted for ADSL also did not support a premature differentiation phenotype (Figure 1—figure supplement 2K), but we recognize that RPE-1 cells may not respond to purine depletion in the same way as neural progenitors or other cell types in vivo.

In RPE-1 and chicken embryos, we observed cell cycle delays following ADSL depletion, as well as an overall reduction in SOX2-positive progenitor cells in both in vivo systems. As a result, differentiated ELAVL3/4-positive cell numbers were also reduced in both chicken and fish systems. Cell cycle delay was further supported by the observation that, contrary to overall Sox2-positive cell numbers, the number of transfected, Sox2-positive cells in chicken embryos was increased relative to controls. In agreement with our findings, nutrient restriction was shown to arrest the proliferation of neural progenitors of Xenopus larvae and zebrafish reversibly in G2, suggesting that most of the cells were quiescent (McKeown and Cline, 2019), consistent with the lack of evidence for cell death or senescence in ADSL-depleted systems. While arrest in Xenopus progenitors did not require mTOR signaling, nutrient-dependent cell cycle reentry was mTOR dependent. Purine deficiency inhibits the mTORC1 pathway, which regulates protein synthesis in response to nutrient availability (Hoxhaj et al., 2017), and inactivation of mTORC1 results in microcephaly in mouse models, but this was attributed mainly to increased levels of cell death (Cloëtta et al., 2013). Thus, while we cannot rule out a role for dysregulated mTOR signaling in the phenotypes associated with ADSLD, the origin of microcephaly is likely to be mechanistically distinct to that in mTORC1-deficient mouse models. We therefore propose that the reduction in brain size resulting from ADSL depletion is largely due to the impaired cell cycle progression observed in SOX2-positive progenitors and delayed differentiation.

The molecular targets of SAICAR remain largely unclear, but its accumulation has been linked to activation of PKM2, and other kinases, in the context of glucose deficiency in cancer (Keller et al., 2012; Keller et al., 2014; Yan et al., 2016). Recent work also implicated ADSL in the activation of MYC, which plays a major role in controlling metabolism and proliferation in cancer cells (Zurlo et al., 2019). Considering that ciliogenesis is a highly regulated process and tightly coordinated with the cell cycle, modulation of central signaling pathways, including AKT and mTOR that control energetics and proliferation, may be one way by which SAICAr could affect cilia (Hoxhaj et al., 2017; Walia et al., 2019). In particular, AKT was shown to regulate a key step in ciliary vesicle formation and AKT activation exerts an inhibitory effect on ciliogenesis induced by serum starvation (Walia et al., 2019). We examined AKT activation following SAICAr treatment and did not observe any detectable increase, indicating that the ability of SAICAr to inhibit ciliogenesis occurs through a distinct mechanism, consistent with the fact that CP110 depletion, which occurs prior to vesicle formation, rescued ciliation in ADSL-depleted cells (Figure 5E). ADSL has also been identified as a proximal interactor of numerous centrosomal proteins, including CEP104, CEP128, CEP135, CEP152, CEP89, and centrobin, suggesting that it may play specific regulatory roles involving direct interactions or its enzymatic activity locally at centrosomes (Gupta et al., 2015). To address this, future work will be needed to determine the cellular interactomes of ADSL and SAICAr, and the signaling they elicit, to identify the disease-relevant targets.

To our knowledge, this is the first demonstration of a specific purine metabolite impairing ciliogenesis. While ciliopathy-like features have not been described for the pathology of ADSLD, we observed a robust rescue of the neural progenitor population in zebrafish following MTX treatment, suggesting that SAICAr and its effects on cilia may be involved. Consistent with effects on cilia in vivo, we observed shorter cilia in the KV of zebrafish, as well as several ciliopathy-related phenotypes (Figures 6A–F ,–7A–G), consistent with impairment of ciliogenesis or cilia function in vivo. Adsl-depleted zebrafish also presented with defects in skull cartilage formation that is coordinated with brain size, in part through cilia-based sonic hedgehog signaling (Xavier et al., 2016). Primary cilia and Hedgehog signaling have well-established roles in regulating multiple progenitor populations in the developing brain (Bangs and Anderson, 2017). However, we also note that severe defects in primary cilia function or Hedgehog signaling cause more drastic reductions in progenitor proliferation and numbers than we observed in either system, potentially consistent with the milder ciliogenesis effects observed in vivo (Breunig et al., 2008; Han et al., 2008). Moreover, we used ADSL knockdown in our experimental systems, and it is currently unclear to what extent the observed ciliary defects would be recapitulated by ADSL mutations in patients and in what tissues and cell types. If ADSL deficiency would be less severe or only a subset of tissues would be affected, patients may not present classic and widespread ciliopathy features. Together, our work provides a cell-level analysis of ADSL deficiency, identifies specific cellular defects, and ascribes these to defects in specific steps of DNPS or the purine nucleotide cycle. Highlighting the complex etiology of ADSLD, our results add further support to the notion that SAICAr may play a key role in pathological outcomes and establish a framework for deciphering the underlying molecular mechanisms.

Summary of phenotypes identified in the systems used. Corresponding figures are indicated, as well as interventions that rescue or recapitulate the phenotype.

Multiple reaction monitoring transitions. See details in Materials and methods.

Most data generated or analysed during this study are included in the manuscript and supporting source data files. Additional source data is available via Figshare, https://doi.org/10.25452/figshare.plus.c.5793614.

1. 1. Stracker T
   2. Lüders J
   3. Dutto I
   4. Philipp M
   5. Pons S

   (2022) Figshare

   Source data supporting 'Pathway specific effects of ADSL deficiency on neurodevelopment'.

   https://doi.org/10.25452/figshare.plus.c.5793614

1. 1. Agarwal S
   2. Chakravarthi B
   3. Behring M
   4. Kim HG
   5. Chandrashekar DS
   6. Gupta N
   7. Bajpai P
   8. Elkholy A
   9. Balasubramanya SAH
   10. Hardy C
   11. Diffalha SA
   12. Varambally S
   13. Manne U

   (2020) PAICS, a Purine Nucleotide Metabolic Enzyme, is Involved in Tumor Growth and the Metastasis of Colorectal Cancer

   Cancers 12:E772.

   https://doi.org/10.3390/cancers12040772

   • PubMed
   • Google Scholar
2. 1. Al-Dosari MS
   2. Shaheen R
   3. Colak D
   4. Alkuraya FS

   (2010) Novel CENPJ mutation causes Seckel syndrome

   Journal of Medical Genetics 47:411–414.

   https://doi.org/10.1136/jmg.2009.076646

   • PubMed
   • Google Scholar
3. 1. Bangs F
   2. Anderson KV

   (2017) Primary Cilia and Mammalian Hedgehog Signaling

   Cold Spring Harbor Perspectives in Biology 9:a028175.

   https://doi.org/10.1101/cshperspect.a028175

   • PubMed
   • Google Scholar
4. 1. Baresova V
   2. Krijt M
   3. Skopova V
   4. Souckova O
   5. Kmoch S
   6. Zikanova M

   (2016) CRISPR-Cas9 induced mutations along de novo purine synthesis in HeLa cells result in accumulation of individual enzyme substrates and affect purinosome formation

   Molecular Genetics and Metabolism 119:270–277.

   https://doi.org/10.1016/j.ymgme.2016.08.004

   • PubMed
   • Google Scholar
5. 1. Bazzi H
   2. Anderson KV

   (2014) Acentriolar mitosis activates a p53-dependent apoptosis pathway in the mouse embryo

   PNAS 111:E1491–E1500.

   https://doi.org/10.1073/pnas.1400568111

   • PubMed
   • Google Scholar
6. 1. Benedict B
   2. van Harn T
   3. Dekker M
   4. Hermsen S
   5. Kucukosmanoglu A
   6. Pieters W
   7. Delzenne-Goette E
   8. Dorsman JC
   9. Petermann E
   10. Foijer F
   11. Te Riele H

   (2018) Loss of p53 suppresses replication-stress-induced DNA breakage in G1/S checkpoint deficient cells

   eLife 7:e37868.

   https://doi.org/10.7554/eLife.37868

   • PubMed
   • Google Scholar
7. 1. Breunig JJ
   2. Sarkisian MR
   3. Arellano JI
   4. Morozov YM
   5. Ayoub AE
   6. Sojitra S
   7. Wang B
   8. Flavell RA
   9. Rakic P
   10. Town T

   (2008) Primary cilia regulate hippocampal neurogenesis by mediating sonic hedgehog signaling

   PNAS 105:13127–13132.

   https://doi.org/10.1073/pnas.0804558105

   • PubMed
   • Google Scholar
8. 1. Burkhalter MD
   2. Fralish GB
   3. Premont RT
   4. Caron MG
   5. Philipp M

   (2013) Grk5l controls heart development by limiting mTOR signaling during symmetry breaking

   Cell Reports 4:625–632.

   https://doi.org/10.1016/j.celrep.2013.07.036

   • PubMed
   • Google Scholar
9. 1. Campeau E
   2. Ruhl VE
   3. Rodier F
   4. Smith CL
   5. Rahmberg BL
   6. Fuss JO
   7. Campisi J
   8. Yaswen P
   9. Cooper PK
   10. Kaufman PD

   (2009) A versatile viral system for expression and depletion of proteins in mammalian cells

   PLOS ONE 4:e6529.

   https://doi.org/10.1371/journal.pone.0006529

   • PubMed
   • Google Scholar
10. 1. Chen P
    2. Wang D
    3. Chen H
    4. Zhou Z
    5. He X

    (2016) The nonessentiality of essential genes in yeast provides therapeutic insights into a human disease

    Genome Research 26:1355–1362.

    https://doi.org/10.1101/gr.205955.116

    • PubMed
    • Google Scholar
11. 1. Cloëtta D
    2. Thomanetz V
    3. Baranek C
    4. Lustenberger RM
    5. Lin S
    6. Oliveri F
    7. Atanasoski S
    8. Rüegg MA

    (2013) Inactivation of mTORC1 in the developing brain causes microcephaly and affects gliogenesis

    The Journal of Neuroscience 33:7799–7810.

    https://doi.org/10.1523/JNEUROSCI.3294-12.2013

    • PubMed
    • Google Scholar
12. 1. Daignan-Fornier B
    2. Pinson B

    (2019) Yeast to Study Human Purine Metabolism Diseases

    Cells 8:E67.

    https://doi.org/10.3390/cells8010067

    • PubMed
    • Google Scholar
13. 1. Dull T
    2. Zufferey R
    3. Kelly M
    4. Mandel RJ
    5. Nguyen M
    6. Trono D
    7. Naldini L

    (1998) A third-generation lentivirus vector with a conditional packaging system

    Journal of Virology 72:8463–8471.

    https://doi.org/10.1128/JVI.72.11.8463-8471.1998

    • PubMed
    • Google Scholar
14. 1. Farooq M
    2. Lindbæk L
    3. Krogh N
    4. Doganli C
    5. Keller C
    6. Mönnich M
    7. Gonçalves AB
    8. Sakthivel S
    9. Mang Y
    10. Fatima A
    11. Andersen VS
    12. Hussain MS
    13. Eiberg H
    14. Hansen L
    15. Kjaer KW
    16. Gopalakrishnan J
    17. Pedersen LB
    18. Møllgård K
    19. Nielsen H
    20. Baig SM
    21. Tommerup N
    22. Christensen ST
    23. Larsen LA

    (2020) RRP7A links primary microcephaly to dysfunction of ribosome biogenesis, resorption of primary cilia, and neurogenesis

    Nature Communications 11:5816.

    https://doi.org/10.1038/s41467-020-19658-0

    • PubMed
    • Google Scholar
15. Preprint

    1. Fenton AR
    2. Janowitz HN
    3. McReynolds MR
    4. Wang W
    5. Hanna-Rose W

    (2017) A Caenorhabditis elegans Model of Adenylosuccinate Lyase Deficiency Reveals Neuromuscular and Reproductive Phenotypes of Distinct Etiology

    bioRxiv.

    https://doi.org/10.1101/181719

    • Google Scholar
16. 1. Fong CS
    2. Mazo G
    3. Das T
    4. Goodman J
    5. Kim M
    6. O’Rourke BP
    7. Izquierdo D
    8. Tsou M-FB

    (2016) 53BP1 and USP28 mediate p53-dependent cell cycle arrest in response to centrosome loss and prolonged mitosis

    eLife 5:e16270.

    https://doi.org/10.7554/eLife.16270

    • PubMed
    • Google Scholar
17. 1. Gabriel E
    2. Wason A
    3. Ramani A
    4. Gooi LM
    5. Keller P
    6. Pozniakovsky A
    7. Poser I
    8. Noack F
    9. Telugu NS
    10. Calegari F
    11. Šarić T
    12. Hescheler J
    13. Hyman AA
    14. Gottardo M
    15. Callaini G
    16. Alkuraya FS
    17. Gopalakrishnan J

    (2016) CPAP promotes timely cilium disassembly to maintain neural progenitor pool

    The EMBO Journal 35:803–819.

    https://doi.org/10.15252/embj.201593679

    • PubMed
    • Google Scholar
18. 1. Giaever G
    2. Chu AM
    3. Ni L
    4. Connelly C
    5. Riles L
    6. Véronneau S
    7. Dow S
    8. Lucau-Danila A
    9. Anderson K
    10. André B
    11. Arkin AP
    12. Astromoff A
    13. El-Bakkoury M
    14. Bangham R
    15. Benito R
    16. Brachat S
    17. Campanaro S
    18. Curtiss M
    19. Davis K
    20. Deutschbauer A
    21. Entian K-D
    22. Flaherty P
    23. Foury F
    24. Garfinkel DJ
    25. Gerstein M
    26. Gotte D
    27. Güldener U
    28. Hegemann JH
    29. Hempel S
    30. Herman Z
    31. Jaramillo DF
    32. Kelly DE
    33. Kelly SL
    34. Kötter P
    35. LaBonte D
    36. Lamb DC
    37. Lan N
    38. Liang H
    39. Liao H
    40. Liu L
    41. Luo C
    42. Lussier M
    43. Mao R
    44. Menard P
    45. Ooi SL
    46. Revuelta JL
    47. Roberts CJ
    48. Rose M
    49. Ross-Macdonald P
    50. Scherens B
    51. Schimmack G
    52. Shafer B
    53. Shoemaker DD
    54. Sookhai-Mahadeo S
    55. Storms RK
    56. Strathern JN
    57. Valle G
    58. Voet M
    59. Volckaert G
    60. Wang C
    61. Ward TR
    62. Wilhelmy J
    63. Winzeler EA
    64. Yang Y
    65. Yen G
    66. Youngman E
    67. Yu K
    68. Bussey H
    69. Boeke JD
    70. Snyder M
    71. Philippsen P
    72. Davis RW
    73. Johnston M

    (2002) Functional profiling of the Saccharomyces cerevisiae genome

    Nature 418:387–391.

    https://doi.org/10.1038/nature00935

    • PubMed
    • Google Scholar
19. 1. Gottifredi V
    2. Shieh S
    3. Taya Y
    4. Prives C

    (2001) p53 accumulates but is functionally impaired when DNA synthesis is blocked

    PNAS 98:1036–1041.

    https://doi.org/10.1073/pnas.021282898

    • PubMed
    • Google Scholar
20. 1. Gupta GD
    2. Coyaud É
    3. Gonçalves J
    4. Mojarad BA
    5. Liu Y
    6. Wu Q
    7. Gheiratmand L
    8. Comartin D
    9. Tkach JM
    10. Cheung SWT
    11. Bashkurov M
    12. Hasegan M
    13. Knight JD
    14. Lin ZY
    15. Schueler M
    16. Hildebrandt F
    17. Moffat J
    18. Gingras AC
    19. Raught B
    20. Pelletier L

    (2015) A Dynamic Protein Interaction Landscape of the Human Centrosome-Cilium Interface

    Cell 163:1484–1499.

    https://doi.org/10.1016/j.cell.2015.10.065

    • PubMed
    • Google Scholar
21. 1. Hamburger V
    2. Hamilton HL

    (1951) A series of normal stages in the development of the chick embryo

    Journal of Morphology 88:49–92.

    https://doi.org/10.1002/aja.1001950404

    • PubMed
    • Google Scholar
22. 1. Han YG
    2. Spassky N
    3. Romaguera-Ros M
    4. Garcia-Verdugo JM
    5. Aguilar A
    6. Schneider-Maunoury S
    7. Alvarez-Buylla A

    (2008) Hedgehog signaling and primary cilia are required for the formation of adult neural stem cells

    Nature Neuroscience 11:277–284.

    https://doi.org/10.1038/nn2059

    • PubMed
    • Google Scholar
23. 1. Herrera A
    2. Saade M
    3. Menendez A
    4. Marti E
    5. Pons S

    (2014) Sustained Wnt/β-catenin signalling causes neuroepithelial aberrations through the accumulation of aPKC at the apical pole

    Nature Communications 5:4168.

    https://doi.org/10.1038/ncomms5168

    • PubMed
    • Google Scholar
24. 1. Holzer K
    2. Drucker E
    3. Roessler S
    4. Dauch D
    5. Heinzmann F
    6. Waldburger N
    7. Eiteneuer EM
    8. Herpel E
    9. Breuhahn K
    10. Zender L
    11. Schirmacher P
    12. Ori A
    13. Singer S

    (2017) Proteomic Analysis Reveals GMP Synthetase as p53 Repression Target in Liver Cancer

    The American Journal of Pathology 187:228–235.

    https://doi.org/10.1016/j.ajpath.2016.09.022

    • PubMed
    • Google Scholar
25. 1. Hoxhaj G
    2. Hughes-Hallett J
    3. Timson RC
    4. Ilagan E
    5. Yuan M
    6. Asara JM
    7. Ben-Sahra I
    8. Manning BD

    (2017) The mTORC1 Signaling Network Senses Changes in Cellular Purine Nucleotide Levels

    Cell Reports 21:1331–1346.

    https://doi.org/10.1016/j.celrep.2017.10.029

    • PubMed
    • Google Scholar
26. 1. Insolera R
    2. Bazzi H
    3. Shao W
    4. Anderson KV
    5. Shi SH

    (2014) Cortical neurogenesis in the absence of centrioles

    Nature Neuroscience 17:1528–1535.

    https://doi.org/10.1038/nn.3831

    • PubMed
    • Google Scholar
27. 1. Jaeken J
    2. Van den Berghe G

    (1984) An infantile autistic syndrome characterised by the presence of succinylpurines in body fluids

    Lancet (London, England) 2:1058–1061.

    https://doi.org/10.1016/S0140-6736(84)91505-8

    • PubMed
    • Google Scholar
28. 1. Jaffe KM
    2. Thiberge SY
    3. Bisher ME
    4. Burdine RD

    (2010) Imaging cilia in zebrafish

    Methods in Cell Biology 97:415–435.

    https://doi.org/10.1016/S0091-679X(10)97022-2

    • PubMed
    • Google Scholar
29. 1. Jurecka A
    2. Zikanova M
    3. Tylki-Szymanska A
    4. Krijt J
    5. Bogdanska A
    6. Gradowska W
    7. Mullerova K
    8. Sykut-Cegielska J
    9. Kmoch S
    10. Pronicka E

    (2008) Clinical, biochemical and molecular findings in seven Polish patients with adenylosuccinate lyase deficiency

    Molecular Genetics and Metabolism 94:435–442.

    https://doi.org/10.1016/j.ymgme.2008.04.013

    • PubMed
    • Google Scholar
30. 1. Jurecka A
    2. Jurkiewicz E
    3. Tylki-Szymanska A

    (2012) Magnetic resonance imaging of the brain in adenylosuccinate lyase deficiency: a report of seven cases and a review of the literature

    European Journal of Pediatrics 171:131–138.

    https://doi.org/10.1007/s00431-011-1503-9

    • PubMed
    • Google Scholar
31. 1. Jurecka A
    2. Zikanova M
    3. Kmoch S
    4. Tylki-Szymańska A

    (2015) Adenylosuccinate lyase deficiency

    Journal of Inherited Metabolic Disease 38:231–242.

    https://doi.org/10.1007/s10545-014-9755-y

    • PubMed
    • Google Scholar
32. 1. Keller KE
    2. Tan IS
    3. Lee YS

    (2012) SAICAR stimulates pyruvate kinase isoform M2 and promotes cancer cell survival in glucose-limited conditions

    Science (New York, N.Y.) 338:1069–1072.

    https://doi.org/10.1126/science.1224409

    • PubMed
    • Google Scholar
33. 1. Keller KE
    2. Doctor ZM
    3. Dwyer ZW
    4. Lee YS

    (2014) SAICAR induces protein kinase activity of PKM2 that is necessary for sustained proliferative signaling of cancer cells

    Molecular Cell 53:700–709.

    https://doi.org/10.1016/j.molcel.2014.02.015

    • PubMed
    • Google Scholar
34. 1. Kim MP
    2. Zhang Y
    3. Lozano G

    (2015) Mutant p53: Multiple Mechanisms Define Biologic Activity in Cancer

    Frontiers in Oncology 5:249.

    https://doi.org/10.3389/fonc.2015.00249

    • PubMed
    • Google Scholar
35. 1. Lambrus BG
    2. Daggubati V
    3. Uetake Y
    4. Scott PM
    5. Clutario KM
    6. Sluder G
    7. Holland AJ

    (2016) A USP28-53BP1-p53-p21 signaling axis arrests growth after centrosome loss or prolonged mitosis

    The Journal of Cell Biology 214:143–153.

    https://doi.org/10.1083/jcb.201604054

    • PubMed
    • Google Scholar
36. 1. Lin YN
    2. Lee YS
    3. Li SK
    4. Tang TK

    (2020) Loss of CPAP in developing mouse brain and its functional implication for human primary microcephaly

    Journal of Cell Science 133:jcs243592.

    https://doi.org/10.1242/jcs.243592

    • PubMed
    • Google Scholar
37. 1. Mádrová L
    2. Krijt M
    3. Barešová V
    4. Václavík J
    5. Friedecký D
    6. Dobešová D
    7. Součková O
    8. Škopová V
    9. Adam T
    10. Zikánová M

    (2018) Mass spectrometric analysis of purine de novo biosynthesis intermediates

    PLOS ONE 13:e0208947.

    https://doi.org/10.1371/journal.pone.0208947

    • PubMed
    • Google Scholar
38. 1. Marjanović M
    2. Sánchez-Huertas C
    3. Terré B
    4. Gómez R
    5. Scheel JF
    6. Pacheco S
    7. Knobel PA
    8. Martínez-Marchal A
    9. Aivio S
    10. Palenzuela L
    11. Wolfrum U
    12. McKinnon PJ
    13. Suja JA
    14. Roig I
    15. Costanzo V
    16. Lüders J
    17. Stracker TH

    (2015) CEP63 deficiency promotes p53-dependent microcephaly and reveals a role for the centrosome in meiotic recombination

    Nature Communications 6:7676.

    https://doi.org/10.1038/ncomms8676

    • PubMed
    • Google Scholar
39. 1. Marsac R
    2. Pinson B
    3. Saint-Marc C
    4. Olmedo M
    5. Artal-Sanz M
    6. Daignan-Fornier B
    7. Gomes JE

    (2019) Purine Homeostasis Is Necessary for Developmental Timing, Germline Maintenance and Muscle Integrity in Caenorhabditis elegans

    Genetics 211:1297–1313.

    https://doi.org/10.1534/genetics.118.301062

    • PubMed
    • Google Scholar
40. 1. McIntyre RE
    2. Lakshminarasimhan Chavali P
    3. Ismail O
    4. Carragher DM
    5. Sanchez-Andrade G
    6. Forment JV
    7. Fu B
    8. Del Castillo Velasco-Herrera M
    9. Edwards A
    10. van der Weyden L
    11. Yang F
    12. Ramirez-Solis R
    13. Estabel J
    14. Gallagher FA
    15. Logan DW
    16. Arends MJ
    17. Tsang SH
    18. Mahajan VB
    19. Scudamore CL
    20. White JK
    21. Jackson SP
    22. Gergely F
    23. Adams DJ
    24. Sanger Mouse Genetics Project

    (2012) Disruption of mouse Cenpj, a regulator of centriole biogenesis, phenocopies Seckel syndrome

    PLOS Genetics 8:e1003022.

    https://doi.org/10.1371/journal.pgen.1003022

    • PubMed
    • Google Scholar
41. 1. McKeown CR
    2. Cline HT

    (2019) Nutrient restriction causes reversible G2 arrest in Xenopus neural progenitors

    Development (Cambridge, England) 146:dev178871.

    https://doi.org/10.1242/dev.178871

    • PubMed
    • Google Scholar
42. 1. Mouchegh K
    2. Zikánová M
    3. Hoffmann GF
    4. Kretzschmar B
    5. Kühn T
    6. Mildenberger E
    7. Stoltenburg-Didinger G
    8. Krijt J
    9. Dvoráková L
    10. Honzík T
    11. Zeman J
    12. Kmoch S
    13. Rossi R

    (2007) Lethal fetal and early neonatal presentation of adenylosuccinate lyase deficiency: observation of 6 patients in 4 families

    The Journal of Pediatrics 150:57–61.

    https://doi.org/10.1016/j.jpeds.2006.09.027

    • PubMed
    • Google Scholar
43. 1. Murga M
    2. Bunting S
    3. Montaña MF
    4. Soria R
    5. Mulero F
    6. Cañamero M
    7. Lee Y
    8. McKinnon PJ
    9. Nussenzweig A
    10. Fernandez-Capetillo O

    (2009) A mouse model of ATR-Seckel shows embryonic replicative stress and accelerated aging

    Nature Genetics 41:891–898.

    https://doi.org/10.1038/ng.420

    • PubMed
    • Google Scholar
44. 1. Ogungbenro YA
    2. Tena TC
    3. Gaboriau D
    4. Lalor P
    5. Dockery P
    6. Philipp M
    7. Morrison CG

    (2018) Centrobin controls primary ciliogenesis in vertebrates

    The Journal of Cell Biology 217:1205–1215.

    https://doi.org/10.1083/jcb.201706095

    • PubMed
    • Google Scholar
45. 1. O’Driscoll M
    2. Ruiz-Perez VL
    3. Woods CG
    4. Jeggo PA
    5. Goodship JA

    (2003) A splicing mutation affecting expression of ataxia-telangiectasia and Rad3-related protein (ATR) results in Seckel syndrome

    Nature Genetics 33:497–501.

    https://doi.org/10.1038/ng1129

    • PubMed
    • Google Scholar
46. 1. Pantoja C
    2. Serrano M

    (1999) Murine fibroblasts lacking p21 undergo senescence and are resistant to transformation by oncogenic Ras

    Oncogene 18:4974–4982.

    https://doi.org/10.1038/sj.onc.1202880

    • PubMed
    • Google Scholar
47. 1. Pinson B
    2. Vaur S
    3. Sagot I
    4. Coulpier F
    5. Lemoine S
    6. Daignan-Fornier B

    (2009) Metabolic intermediates selectively stimulate transcription factor interaction and modulate phosphate and purine pathways

    Genes & Development 23:1399–1407.

    https://doi.org/10.1101/gad.521809

    • PubMed
    • Google Scholar
48. 1. Reddy BA
    2. van der Knaap JA
    3. Bot AGM
    4. Mohd-Sarip A
    5. Dekkers DHW
    6. Timmermans MA
    7. Martens JWM
    8. Demmers JAA
    9. Verrijzer CP

    (2014) Nucleotide biosynthetic enzyme GMP synthase is a TRIM21-controlled relay of p53 stabilization

    Molecular Cell 53:458–470.

    https://doi.org/10.1016/j.molcel.2013.12.017

    • PubMed
    • Google Scholar
49. 1. Sir J-H
    2. Barr AR
    3. Nicholas AK
    4. Carvalho OP
    5. Khurshid M
    6. Sossick A
    7. Reichelt S
    8. D’Santos C
    9. Woods CG
    10. Gergely F

    (2011) A primary microcephaly protein complex forms a ring around parental centrioles

    Nature Genetics 43:1147–1153.

    https://doi.org/10.1038/ng.971

    • PubMed
    • Google Scholar
50. 1. Stewart SA
    2. Dykxhoorn DM
    3. Palliser D
    4. Mizuno H
    5. Yu EY
    6. An DS
    7. Sabatini DM
    8. Chen ISY
    9. Hahn WC
    10. Sharp PA
    11. Weinberg RA
    12. Novina CD

    (2003) Lentivirus-delivered stable gene silencing by RNAi in primary cells

    RNA (New York, N.Y.) 9:493–501.

    https://doi.org/10.1261/rna.2192803

    • PubMed
    • Google Scholar
51. 1. Stone TW
    2. Roberts LA
    3. Morris BJ
    4. Jones PA
    5. Ogilvy HA
    6. Behan WM
    7. Duley JA
    8. Simmonds HA
    9. Vincent MF
    10. van den Berghe G

    (1998) Succinylpurines induce neuronal damage in the rat brain

    Advances in Experimental Medicine and Biology 431:185–189.

    https://doi.org/10.1007/978-1-4615-5381-6_36

    • PubMed
    • Google Scholar
52. 1. Stracker TH
    2. Morrison CG
    3. Gergely F

    (2020) Molecular causes of primary microcephaly and related diseases: a report from the UNIA Workshop

    Chromosoma 129:115–120.

    https://doi.org/10.1007/s00412-020-00737-6

    • PubMed
    • Google Scholar
53. 1. Taha-Mehlitz S
    2. Bianco G
    3. Coto-Llerena M
    4. Kancherla V
    5. Bantug GR
    6. Gallon J
    7. Ercan C
    8. Panebianco F
    9. Eppenberger-Castori S
    10. von Strauss M
    11. Staubli S
    12. Bolli M
    13. Peterli R
    14. Matter MS
    15. Terracciano LM
    16. von Flüe M
    17. Ng CKY
    18. Soysal SD
    19. Kollmar O
    20. Piscuoglio S

    (2021) Adenylosuccinate lyase is oncogenic in colorectal cancer by causing mitochondrial dysfunction and independent activation of NRF2 and mTOR-MYC-axis

    Theranostics 11:4011–4029.

    https://doi.org/10.7150/thno.50051

    • PubMed
    • Google Scholar
54. 1. Tamiya S
    2. Liu L
    3. Kaplan HJ

    (2010) Epithelial-mesenchymal transition and proliferation of retinal pigment epithelial cells initiated upon loss of cell-cell contact

    Investigative Ophthalmology & Visual Science 51:2755–2763.

    https://doi.org/10.1167/iovs.09-4725

    • PubMed
    • Google Scholar
55. 1. Thisse B
    2. Thisse C

    (2014) In situ hybridization on whole-mount zebrafish embryos and young larvae

    Methods in Molecular Biology (Clifton, N.J.) 1211:53–67.

    https://doi.org/10.1007/978-1-4939-1459-3_5

    • PubMed
    • Google Scholar
56. 1. Vaz R
    2. Hofmeister W
    3. Lindstrand A

    (2019) Zebrafish Models of Neurodevelopmental Disorders: Limitations and Benefits of Current Tools and Techniques

    International Journal of Molecular Sciences 20:E1296.

    https://doi.org/10.3390/ijms20061296

    • PubMed
    • Google Scholar
57. 1. Walia V
    2. Cuenca A
    3. Vetter M
    4. Insinna C
    5. Perera S
    6. Lu Q
    7. Ritt DA
    8. Semler E
    9. Specht S
    10. Stauffer J
    11. Morrison DK
    12. Lorentzen E
    13. Westlake CJ

    (2019) Akt Regulates a Rab11-Effector Switch Required for Ciliogenesis

    Developmental Cell 50:229–246.

    https://doi.org/10.1016/j.devcel.2019.05.022

    • PubMed
    • Google Scholar
58. 1. Xavier GM
    2. Seppala M
    3. Barrell W
    4. Birjandi AA
    5. Geoghegan F
    6. Cobourne MT

    (2016) Hedgehog receptor function during craniofacial development

    Developmental Biology 415:198–215.

    https://doi.org/10.1016/j.ydbio.2016.02.009

    • PubMed
    • Google Scholar
59. 1. Yamauchi T
    2. Miyawaki K
    3. Semba Y
    4. Takahashi M
    5. Izumi Y
    6. Nogami J
    7. Nakao F
    8. Sugio T
    9. Sasaki K
    10. Pinello L
    11. Bauer DE
    12. Bamba T
    13. Akashi K
    14. Maeda T

    (2022) Targeting leukemia-specific dependence on the de novo purine synthesis pathway

    Leukemia 36:383–393.

    https://doi.org/10.1038/s41375-021-01369-0

    • PubMed
    • Google Scholar
60. 1. Yan M
    2. Chakravarthy S
    3. Tokuda JM
    4. Pollack L
    5. Bowman GD
    6. Lee YS

    (2016) Succinyl-5-aminoimidazole-4-carboxamide-1-ribose 5’-Phosphate (SAICAR) Activates Pyruvate Kinase Isoform M2 (PKM2) in Its Dimeric Form

    Biochemistry 55:4731–4736.

    https://doi.org/10.1021/acs.biochem.6b00658

    • PubMed
    • Google Scholar
61. 1. Zikánová M
    2. Krijt J
    3. Hartmannová H
    4. Kmoch S

    (2005) Preparation of 5-amino-4-imidazole-N-succinocarboxamide ribotide, 5-amino-4-imidazole-N-succinocarboxamide riboside and succinyladenosine, compounds usable in diagnosis and research of adenylosuccinate lyase deficiency

    Journal of Inherited Metabolic Disease 28:493–499.

    https://doi.org/10.1007/s10545-005-0493-z

    • PubMed
    • Google Scholar
62. 1. Zurlo G
    2. Liu X
    3. Takada M
    4. Fan C
    5. Simon JM
    6. Ptacek TS
    7. Rodriguez J
    8. von Kriegsheim A
    9. Liu J
    10. Locasale JW
    11. Robinson A
    12. Zhang J
    13. Holler JM
    14. Kim B
    15. Zikánová M
    16. Bierau J
    17. Xie L
    18. Chen X
    19. Li M
    20. Perou CM
    21. Zhang Q

    (2019) Prolyl hydroxylase substrate adenylosuccinate lyase is an oncogenic driver in triple negative breast cancer

    Nature Communications 10:5177.

    https://doi.org/10.1038/s41467-019-13168-4

    • PubMed
    • Google Scholar

Author details

1. Ilaria Dutto

   Institute for Research in Biomedicine, The Barcelona Institute of Science and Technology, Barcelona, Spain

   Contribution

   Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Visualization, Writing – original draft, Writing – review and editing

   Competing interests

   No competing interests declared

2. Julian Gerhards

   1. Department of Experimental and Clinical Pharmacology and Pharmacogenomics, University of Tübingen, Tübingen, Germany
   2. Institute of Biochemistry and Molecular Biology, Ulm University, Ulm, Germany

   Contribution

   Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Visualization, Writing – original draft, Writing – review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-7005-1618

3. Antonio Herrera

   Department of Cell Biology, Instituto de Biología Molecular de Barcelona, Barcelona, Spain

   Contribution

   Data curation, Formal analysis, Investigation, Methodology, Visualization, Writing – original draft, Writing – review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-6248-1001

4. Olga Souckova

   Department of Paediatrics and Inherited Metabolic Disorders, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic

   Contribution

   Formal analysis, Investigation

   Competing interests

   No competing interests declared

5. Václava Škopová

   Department of Paediatrics and Inherited Metabolic Disorders, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic

   Contribution

   Formal analysis, Investigation

   Competing interests

   No competing interests declared

6. Jordann A Smak

   National Cancer Institute, Center for Cancer Research, Radiation Oncology Branch, Bethesda, United States

   Contribution

   Formal analysis, Investigation

   Competing interests

   No competing interests declared

7. Alexandra Junza

   1. Universitat Rovira i Virgili, Department of Electronic Engineering, IISPV, Tarragona, Spain
   2. CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Instituto de Salud Carlos III, Madrid, Spain

   Contribution

   Formal analysis, Investigation

   Competing interests

   No competing interests declared

8. Oscar Yanes

   1. Universitat Rovira i Virgili, Department of Electronic Engineering, IISPV, Tarragona, Spain
   2. CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Instituto de Salud Carlos III, Madrid, Spain

   Contribution

   Data curation, Formal analysis, Investigation, Methodology

   Competing interests

   No competing interests declared

9. Cedric Boeckx

   1. ICREA, Barcelona, Spain
   2. Institute of Complex Systems (UBICS), Universitat de Barcelona, Barcelona, Spain
   3. Section of General Linguistics, Universitat de Barcelona, Barcelona, Spain

   Contribution

   Conceptualization

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-8882-9718

10. Martin D Burkhalter

    Department of Experimental and Clinical Pharmacology and Pharmacogenomics, University of Tübingen, Tübingen, Germany

    Contribution

    Formal analysis, Investigation

    Competing interests

    No competing interests declared

    "This ORCID iD identifies the author of this article:" 0000-0002-8646-3131

11. Marie Zikánová

    Department of Paediatrics and Inherited Metabolic Disorders, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic

    Contribution

    Formal analysis, Investigation, Methodology, Supervision, Writing – review and editing

    Competing interests

    No competing interests declared

12. Sebastian Pons

    Department of Cell Biology, Instituto de Biología Molecular de Barcelona, Barcelona, Spain

    Contribution

    Formal analysis, Methodology, Resources, Supervision

    Competing interests

    No competing interests declared

13. Melanie Philipp

    1. Department of Experimental and Clinical Pharmacology and Pharmacogenomics, University of Tübingen, Tübingen, Germany
    2. Institute of Biochemistry and Molecular Biology, Ulm University, Ulm, Germany

    Contribution

    Data curation, Formal analysis, Funding acquisition, Investigation, Methodology, Supervision, Visualization, Writing – original draft, Writing – review and editing

    Competing interests

    No competing interests declared

    "This ORCID iD identifies the author of this article:" 0000-0003-2714-965X

14. Jens Lüders

    Institute for Research in Biomedicine, The Barcelona Institute of Science and Technology, Barcelona, Spain

    Contribution

    Conceptualization, Data curation, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Visualization, Writing – original draft, Writing – review and editing

    For correspondence

    jens.luders@irbbarcelona.org

    Competing interests

    No competing interests declared

    "This ORCID iD identifies the author of this article:" 0000-0002-9018-7977

15. Travis H Stracker

    1. Institute for Research in Biomedicine, The Barcelona Institute of Science and Technology, Barcelona, Spain
    2. National Cancer Institute, Center for Cancer Research, Radiation Oncology Branch, Bethesda, United States

    Contribution

    Conceptualization, Data curation, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Visualization, Writing – original draft, Writing – review and editing

    For correspondence

    travis.stracker@nih.gov

    Competing interests

    No competing interests declared

    "This ORCID iD identifies the author of this article:" 0000-0002-8650-2081

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