RNA polymerase (pol) III transcribes various untranslated RNAs including 5S rRNA and tRNAs. RNA pol III-derived transcripts play essential roles in several processes, including protein synthesis and secretion (Dieci et al., 2013; Dieci et al., 2007). In addition to RNA pol III, transcription of tRNAs requires the recruitment of TFIIIC and TFIIIB to the promoter. TFIIIB consists of Brf1, TATA-binding protein (TBP) and B-double prime (Orioli et al., 2012). RNA pol III-dependent transcription is also tightly regulated through either direct or indirect mechanisms that control TFIIIB recruitment to the promotor (Gomez-Roman et al., 2003; Kenneth et al., 2007; Felton-Edkins et al., 2003; Sriskanthadevan-Pirahas et al., 2018; Crighton et al., 2003; Sutcliffe et al., 2000; White et al., 1996; Woiwode et al., 2008). MAF1 is a key repressor of RNA pol III-dependent transcription ( Graczyk et al., 2015; Johnson et al., 2007). It binds to RNA pol III and prevents the interaction between RNA pol III and TFIIIB (Vorländer et al., 2019; Vannini et al., 2010). MAF1 has also been shown to regulate a variety of RNA pol II-transcribed targets (Johnson et al., 2007; Palian et al., 2014; Khanna et al., 2014; Li et al., 2016; Lee et al., 2015), and acts as a tumor suppressor (Palian et al., 2014; Li et al., 2016), regulates metabolism (Willis et al., 2018; Bonhoure et al., 2015), and longevity (Shetty et al., 2020; Cai and Wei, 2016).

RNA pol III-mediated transcription has been shown to play an important role during development and cellular differentiation. Two RNA pol III isoforms are differentially expressed between pluripotent and differentiated embryonic stem cells (ESCs) (Haurie et al., 2010; Wong et al., 2011; Wang et al., 2020). RNA pol III-dependent transcription also modulates the formation of hematopoietic lineages in zebrafish (Wei et al., 2016) and transcription is downregulated during skeletal muscle differentiation of Xenopus tropicalis (McQueen et al., 2019). MAF1 enhances the formation of mesoderm in embryonic stem cells, and the downregulation of RNA pol III-dependent transcription enhances the differentiation of ESCs and 3T3-L1 cells into adipocytes (Chen et al., 2018).

Diseases associated with ribosomal disfunctions, ribosomopathies, are commonly associated with bone marrow, skeletal and craniofacial disorders (Trainor and Merrill, 2014). This surprising tissue specificity suggests that these tissues may be particularly sensitive to alterations in protein synthesis. For example, Treacher Collins syndrome is caused by mutations in POLR1C, POLR1D, or TCOF1, which affect rDNA transcription by RNA pol I and ribosome biogenesis (Noack Watt et al., 2016; Dauwerse et al., 2011). RNA pol III-derived transcripts may also play a role as POLR1C and POLR1D are common subunits of both RNA pol I and RNA pol III. In yeast, Treacher Collins syndrome-related mutations in POLR1D result in altered functions of both RNA pol I and III (Walker-Kopp et al., 2017). Furthermore, RNA pol III-dependent transcription may play a role in cerebellofaciodental syndrome which is associated with mutations in the RNA pol III-specific transcription factor, Brf1. This syndrome is characterized by a neurodevelopmental phenotype as well as changes in the facial and dental structure and delayed bone age (Borck et al., 2015; Jee et al., 2017; Honjo et al., 2021). Additionally, bone phenotypes have been described in a subgroup of patients with mutations in RNA pol III subunits (Borck et al., 2015; Jee et al., 2017; Honjo et al., 2021; Terhal et al., 2020; Ghoumid et al., 2017).

It is known that MAF1 regulates mesoderm formation and adipocyte differentiation (Chen et al., 2018). Osteoblasts and adipocytes are both derived from the mesenchymal lineage (Chen et al., 2016) and mutations in BRF1 and RNA pol III subunits are associated with bone-related phenotypes. These facts led us to hypothesize that regulation of RNA pol III-dependent transcription by MAF1 may play a fundamental role in osteoblast differentiation, bone formation and hence, bone mass. Here, we show that both whole body deletion of MAF1 in mice and tissue-specific overexpression of MAF1 in stromal cells of the long bones enhances bone mass in vivo, while MAF1 induces osteoblast differentiation in vitro. To further examine the role of MAF1 and RNA pol III-mediated transcription in osteoblast differentiation, we attempted to study the effect on osteoblast differentiation by repressing transcription through different approaches. Surprisingly, while MAF1 induced osteoblast differentiation, repression of RNA pol III-dependent transcription, by either chemical inhibition of RNA pol III or by Brf1 knockdown, decreased osteoblast differentiation. Thus, changes in MAF1 expression produce an opposing effect on osteoblast differentiation compared with other approaches that repress RNA pol III-mediated transcription. We further show that these three different approaches to decrease RNA pol III-dependent transcription result in divergent gene expression changes. Altered MAF1 expression affects the expression of the osteoblast differentiation gene program. Together, these findings reveal that MAF1 plays a key role in osteoblast differentiation and bone mass regulation and that its ability to regulate RNA pol III-dependent transcription contributes to the observed phenotypic outcomes.

MAF1 is a key repressor of transcription by RNA pol III (Johnson et al., 2007; Vorländer et al., 2019). Changes in MAF1 expression have been shown to enhance adipocyte differentiation (Chen et al., 2018; Figure 2—figure supplement 1 and Figure 3—figure supplement 1). Maf1^-/- mice are shorter than WT mice and exhibit a lean phenotype with resistance to diet-induced obesity, decreased fertility and fecundity, and increased longevity/healthspan (Bonhoure et al., 2015). Here, we demonstrate that these mice display an increase in bone volume and bone formation. However, Maf1^-/- derived primary stromal cells showed a decrease in osteoblast formation in ex vivo cultures. While it is possible this is due to different culture conditions, this latter finding was consistent with the effects of MAF1 overexpression specifically in the mesenchyme of long bones, which resulted in enhanced osteoblast differentiation and an increase in bone mass. In these latter mice, compared with Maf1^-/- mice, any confounding actions due to the absence of MAF1 in other tissues, such as possible endocrine or paracrine effects, are limited. Thus, our results indicate that the ability of MAF1 to regulate bone mass involves both cell autonomous and non-cell-autonomous actions. This idea is further corroborated by our in vitro results showing that osteoblast differentiation is regulated by increasing or decreasing MAF1 expression in ST2 cells, suggesting that MAF1 promotes osteoblast differentiation and mineralization. Our findings are summarized in Table 1. As MAF1 also enhances adipogenesis (Chen et al., 2018; Figure 2—figure supplement 1, Figure 3—figure supplement 1), we conclude that MAF1 is an important regulator in the development and differentiation of mesenchymal cells into multiple lineages.

MAF1 is a well-established repressor of RNA pol III-mediated transcription through its direct interaction with RNA pol III (Vorländer et al., 2019; Vannini et al., 2010). To further determine whether MAF1 functions to promote osteoblast differentiation through its ability to regulate RNA pol III-dependent transcription, we used complementary approaches to repress this transcription process. During differentiation, we observed an overall increase in tRNA gene transcripts. This increase was repressed by MAF1 overexpression, Brf1 downregulation, or chemical inhibition of RNA pol III. Surprisingly, however, in contrast to the positive regulation of osteoblast differentiation by MAF1, chemical inhibition of RNA pol III or Brf1 knockdown resulted in a decrease in osteoblast differentiation (Table 1). Thus, while different perturbations in RNA pol III-dependent transcription all affect the differentiation process, MAF1-mediated changes produce an opposing action compared with chemical inhibition of RNA pol III or decreased Brf1 expression. This is in contrast to what we observed for adipogenesis, where all three approaches to repress RNA pol III transcription similarly increased adipocyte formation (Figure 2—figure supplement 1, Figure 3—figure supplement 1, Figure 4—figure supplement 2, Figure 5—figure supplement 1 and Table 1; Chen et al., 2018).

To understand the basis of the different osteoblast differentiation outcomes, we examined changes in gene expression that resulted from the different perturbations of RNA pol III-dependent transcription. RNA seq revealed that alterations in RNA pol III-mediated transcription prior to, and during differentiation, result in a limited number of overlapping changes, with the four conditions largely producing distinct changes in gene expression profiles. Changes in several established regulators of osteoblast formation and function were identified that correlated with the differentiation outcomes. Overall, the different gene expression profiles likely contribute to the differences in osteoblast differentiation that we observe. However, the precise mechanism underlying the difference between MAF1-regulated changes compared with the alternate approaches to repress RNA pol III-mediated transcription remains unclear. Unlike RNA pol III and Brf1, MAF1 is not an essential RNA pol III transcription factor, and it functions to repress this transcription process by directly interacting with RNA pol III (Vorländer et al., 2019). Given that MAF1 is recruited to certain RNA pol II promoters (Johnson et al., 2007; Palian et al., 2014; Khanna et al., 2014; Li et al., 2016), it is conceivable that the ability of MAF1 to regulate both RNA pol III- and RNA pol II-transcribed genes contributes to its ability to drive osteoblast differentiation. This may also explain its differential effect on osteoblast differentiation when compared with the alternative approaches used to selectively repress RNA pol III-dependent transcription. However, RNA seq did not detect changes in genes previously reported to be regulated by MAF1, such as Tbp and Fasn. The same finding was obtained in an RNA-seq analysis in Maf1^-/- liver (Bonhoure et al., 2020). Thus, the regulation of RNA pol II-dependent gene expression by MAF1 is likely context dependent. Additional studies suggest that the regulation of RNA pol II targets by MAF1 is limited (Orioli et al., 2016). Therefore, we cannot exclude a possible role for MAF1 regulation of RNA pol II transcription in the regulation of osteoblast differentiation. However, since other approaches used to modulate RNA pol III-mediated transcription also alter osteoblast differentiation, MAF1 functions, at least in part, to regulate osteoblastogenesis through its effect on RNA pol III.

In addition to different effects on gene expression by manipulation of MAF1 expression, RNA-seq analysis revealed distinct changes in gene expression exhibited by chemical inhibition of RNA pol III and Brf1 knockdown, despite similar outcomes on osteoblast differentiation. These data indicate that manipulating RNA pol III transcription using different approaches results in disparate outcomes on gene expression. This could be due to differential changes in the tRNA population. The prevalence of specific tRNAs has been correlated with codon-biased translation in multiple tissues (Dittmar et al., 2006; Gobet et al., 2020; Kutter et al., 2011; Rak et al., 2018). Our analysis of mRNA changes during osteoblast differentiation showed that a significant codon bias emerges during this process. Thus, it is conceivable that during osteoblast differentiation specific changes in tRNAs are required to efficiently drive codon-biased translation of mRNAs needed for differentiation to proceed. Thus, this may be a mechanism in which RNA pol III transcription affects osteoblast differentiation. There are several other mechanisms that could contribute to the observed differences in gene regulation by RNA pol III-mediated transcription. RNA pol III transcribes a variety of untranslated RNAs and changes in any of these RNA pol III-derived transcripts may potentially play a role. Additionally, the generation of tRNA fragments (Schimmel, 2018; Su et al., 2020), or the regulation of RNA pol II genes through the recruitment of RNA pol III to nearby SINE sites such as described for CDKN1a (Lee et al., 2015), could all potentially contribute to the differential expression of osteoblast genes when RNA pol III-mediated transcription is altered. Future work will be needed to identify the specific mechanisms by which MAF1 and RNA pol III-mediated transcription alter gene expression to regulate osteoblast differentiation.

In all, our results describe a novel role for MAF1 and RNA pol III in bone biology. Given that different approaches used to modulate RNA pol III-dependent transcription also affect osteoblast differentiation, albeit in an opposing direction from MAF1-mediated effects, the findings support the idea that MAF1 functions, at least in part, to regulate osteoblast development and bone mass through its ability to control RNA pol III-mediated transcription. Distinct qualitative or quantitative changes resulting from different perturbations in RNA pol III-dependent transcription may also play a role in developmental disorders. Interestingly, several different syndromes relating to mutations in RNA pol III subunits show very heterogeneous phenotypes. This includes POLR3-related hypomyelinating leukodystrophies (Lata et al., 2021; Yeganeh and Hernandez, 2020; Thomas and Thomas, 2019), Wiedemann-Rautenstrauch syndrome, a neonatal progeroid syndrome (Wu et al., 2021b; Wambach et al., 2018; Paolacci et al., 2017; Beauregard-Lacroix et al., 2020), and cerebellar hypoplasia with endosteal sclerosis (Terhal et al., 2020; Ghoumid et al., 2017). Additionally, Brf1 mutations have been shown to be causative for cerebellofaciodental syndrome (Borck et al., 2015; Jee et al., 2017; Honjo et al., 2021; Valenzuela et al., 2020). Some patients with RNA pol III-related mutations, but not all, show bone-related phenotypes (Borck et al., 2015; Jee et al., 2017; Honjo et al., 2021; Terhal et al., 2020; Ghoumid et al., 2017). The considerable heterogeneity of these syndromes has been suggested to be related to differential changes in RNA pol III-dependent transcription (Yeganeh and Hernandez, 2020). Thus, some cells and tissues, including bone and its cells, may be more sensitive to disruption of RNA pol III-mediated transcription. Together, these collective findings and our current study indicate that the exquisite regulation of RNA pol III plays an essential role in a variety of biological and developmental processes.

Sequencing data will be deposited in GEO Source data files will be provided All data will be made available to the public. Raw and processed data from the RNA sequencing experiment determining gene expression before and during osteoblast differentiation has been uploaded to the GEO data base with accession nr. GSE203308.

1. 1. Phillips E
   2. Ahmad N
   3. Sun L
   4. Iben J
   5. Walkey CJ
   6. Rusin A
   7. Yuen T
   8. Rosen CJ
   9. Willis IM
   10. Zaidi M
   11. Johnson DL

   (2022) NCBI Gene Expression Omnibus

   ID GSE203308. RNA seq after RNA pol III manipulation by Brf1 and Maf1 knockdown, Maf1 overexpression and ML60218 treatment in ST2 cells before, and during differentiation into osteoblasts.

   https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE203308

1. 1. Beauregard-Lacroix E
   2. Salian S
   3. Kim H
   4. Ehresmann S
   5. DʹAmours G
   6. Gauthier J
   7. Saillour V
   8. Bernard G
   9. Mitchell GA
   10. Soucy J-F
   11. Michaud JL
   12. Campeau PM

   (2020) A variant of neonatal progeroid syndrome, or Wiedemann-Rautenstrauch syndrome, is associated with A nonsense variant in POLR3GL

   European Journal of Human Genetics 28:461–468.

   https://doi.org/10.1038/s41431-019-0539-6

   • PubMed
   • Google Scholar
2. 1. Bonhoure N
   2. Byrnes A
   3. Moir RD
   4. Hodroj W
   5. Preitner F
   6. Praz V
   7. Marcelin G
   8. Chua SC Jr
   9. Martinez-Lopez N
   10. Singh R
   11. Moullan N
   12. Auwerx J
   13. Willemin G
   14. Shah H
   15. Hartil K
   16. Vaitheesvaran B
   17. Kurland I
   18. Hernandez N
   19. Willis IM

   (2015) Loss of the RNA polymerase III repressor MAF1 confers obesity resistance

   Genes & Development 29:934–947.

   https://doi.org/10.1101/gad.258350.115

   • PubMed
   • Google Scholar
3. 1. Bonhoure N
   2. Praz V
   3. Moir RD
   4. Willemin G
   5. Mange F
   6. Moret C
   7. Willis IM
   8. Hernandez N

   (2020) MAF1 is a chronic repressor of RNA polymerase III transcription in the mouse

   Scientific Reports 10:11956.

   https://doi.org/10.1038/s41598-020-68665-0

   • PubMed
   • Google Scholar
4. 1. Borck G
   2. Hög F
   3. Dentici ML
   4. Tan PL
   5. Sowada N
   6. Medeira A
   7. Gueneau L
   8. Thiele H
   9. Kousi M
   10. Lepri F
   11. Wenzeck L
   12. Blumenthal I
   13. Radicioni A
   14. Schwarzenberg TL
   15. Mandriani B
   16. Fischetto R
   17. Morris-Rosendahl DJ
   18. Altmüller J
   19. Reymond A
   20. Nürnberg P
   21. Merla G
   22. Dallapiccola B
   23. Katsanis N
   24. Cramer P
   25. Kubisch C

   (2015) BRF1 mutations alter RNA polymerase III-dependent transcription and cause neurodevelopmental anomalies

   Genome Research 25:155–166.

   https://doi.org/10.1101/gr.176925.114

   • PubMed
   • Google Scholar
5. 1. Cai Y
   2. Wei Y-H

   (2016) Stress resistance and lifespan are increased in C. elegans but decreased in S. cerevisiae by mafr-1/maf1 deletion

   Oncotarget 7:10812–10826.

   https://doi.org/10.18632/oncotarget.7769

   • PubMed
   • Google Scholar
6. 1. Chen Q
   2. Shou P
   3. Zheng C
   4. Jiang M
   5. Cao G
   6. Yang Q
   7. Cao J
   8. Xie N
   9. Velletri T
   10. Zhang X
   11. Xu C
   12. Zhang L
   13. Yang H
   14. Hou J
   15. Wang Y
   16. Shi Y

   (2016) Fate decision of mesenchymal stem cells: adipocytes or osteoblasts?

   Cell Death and Differentiation 23:1128–1139.

   https://doi.org/10.1038/cdd.2015.168

   • PubMed
   • Google Scholar
7. 1. Chen C-Y
   2. Lanz RB
   3. Walkey CJ
   4. Chang W-H
   5. Lu W
   6. Johnson DL

   (2018) Maf1 and Repression of RNA Polymerase III-Mediated Transcription Drive Adipocyte Differentiation

   Cell Reports 24:1852–1864.

   https://doi.org/10.1016/j.celrep.2018.07.046

   • PubMed
   • Google Scholar
8. 1. Crighton D
   2. Woiwode A
   3. Zhang C
   4. Mandavia N
   5. Morton JP
   6. Warnock LJ
   7. Milner J
   8. White RJ
   9. Johnson DL

   (2003) p53 represses RNA polymerase III transcription by targeting TBP and inhibiting promoter occupancy by TFIIIB

   The EMBO Journal 22:2810–2820.

   https://doi.org/10.1093/emboj/cdg265

   • PubMed
   • Google Scholar
9. 1. Dauwerse JG
   2. Dixon J
   3. Seland S
   4. Ruivenkamp CAL
   5. van Haeringen A
   6. Hoefsloot LH
   7. Peters DJM
   8. Boers AC
   9. Daumer-Haas C
   10. Maiwald R
   11. Zweier C
   12. Kerr B
   13. Cobo AM
   14. Toral JF
   15. Hoogeboom AJM
   16. Lohmann DR
   17. Hehr U
   18. Dixon MJ
   19. Breuning MH
   20. Wieczorek D

   (2011) Mutations in genes encoding subunits of RNA polymerases I and III cause Treacher Collins syndrome

   Nature Genetics 43:20–22.

   https://doi.org/10.1038/ng.724

   • PubMed
   • Google Scholar
10. 1. Dieci G
    2. Fiorino G
    3. Castelnuovo M
    4. Teichmann M
    5. Pagano A

    (2007) The expanding RNA polymerase III transcriptome

    Trends in Genetics 23:614–622.

    https://doi.org/10.1016/j.tig.2007.09.001

    • PubMed
    • Google Scholar
11. 1. Dieci G
    2. Conti A
    3. Pagano A
    4. Carnevali D

    (2013) Identification of RNA polymerase III-transcribed genes in eukaryotic genomes

    Biochimica et Biophysica Acta 1829:296–305.

    https://doi.org/10.1016/j.bbagrm.2012.09.010

    • PubMed
    • Google Scholar
12. 1. Dittmar KA
    2. Goodenbour JM
    3. Pan T

    (2006) Tissue-specific differences in human transfer RNA expression

    PLOS Genetics 2:e0020221.

    https://doi.org/10.1371/journal.pgen.0020221

    • PubMed
    • Google Scholar
13. 1. Felton-Edkins ZA
    2. Fairley JA
    3. Graham EL
    4. Johnston IM
    5. White RJ
    6. Scott PH

    (2003) The mitogen-activated protein (MAP) kinase ERK induces tRNA synthesis by phosphorylating TFIIIB

    The EMBO Journal 22:2422–2432.

    https://doi.org/10.1093/emboj/cdg240

    • PubMed
    • Google Scholar
14. 1. Fitter S
    2. Matthews MP
    3. Martin SK
    4. Xie J
    5. Ooi SS
    6. Walkley CR
    7. Codrington JD
    8. Ruegg MA
    9. Hall MN
    10. Proud CG
    11. Gronthos S
    12. Zannettino ACW

    (2017) mTORC1 Plays an Important Role in Skeletal Development by Controlling Preosteoblast Differentiation

    Molecular and Cellular Biology 37:1–20.

    https://doi.org/10.1128/MCB.00668-16

    • PubMed
    • Google Scholar
15. 1. Fujioka-Kobayashi M
    2. Caballé-Serrano J
    3. Bosshardt DD
    4. Gruber R
    5. Buser D
    6. Miron RJ

    (2016) Bone conditioned media (BCM) improves osteoblast adhesion and differentiation on collagen barrier membranes

    BMC Oral Health 17:1–7.

    https://doi.org/10.1186/s12903-016-0230-z

    • PubMed
    • Google Scholar
16. 1. Ghoumid J
    2. Petit F
    3. Boute-Benejean O
    4. Frenois F
    5. Cartigny M
    6. Vanlerberghe C
    7. Smol T
    8. Caumes R
    9. de Roux N
    10. Manouvrier-Hanu S

    (2017) Cerebellar hypoplasia with endosteal sclerosis is a POLR3-related disorder

    European Journal of Human Genetics 25:1011–1014.

    https://doi.org/10.1038/ejhg.2017.73

    • PubMed
    • Google Scholar
17. 1. Gobet C
    2. Weger BD
    3. Marquis J
    4. Martin E
    5. Neelagandan N
    6. Gachon F
    7. Naef F

    (2020) Robust landscapes of ribosome dwell times and aminoacyl-tRNAs in response to nutrient stress in liver

    PNAS 117:9630–9641.

    https://doi.org/10.1073/pnas.1918145117

    • PubMed
    • Google Scholar
18. 1. Gomez-Roman N
    2. Grandori C
    3. Eisenman RN
    4. White RJ

    (2003) Direct activation of RNA polymerase III transcription by c-Myc

    Nature 421:290–294.

    https://doi.org/10.1038/nature01327

    • PubMed
    • Google Scholar
19. 1. Graczyk D
    2. White RJ
    3. Ryan KM

    (2015) Involvement of RNA Polymerase III in Immune Responses

    Molecular and Cellular Biology 35:1848–1859.

    https://doi.org/10.1128/MCB.00990-14

    • PubMed
    • Google Scholar
20. 1. Graczyk D
    2. Cieśla M
    3. Boguta M

    (2018) Regulation of tRNA synthesis by the general transcription factors of RNA polymerase III - TFIIIB and TFIIIC, and by the MAF1 protein

    Biochimica et Biophysica Acta. Gene Regulatory Mechanisms 1861:320–329.

    https://doi.org/10.1016/j.bbagrm.2018.01.011

    • PubMed
    • Google Scholar
21. 1. Haurie V
    2. Durrieu-Gaillard S
    3. Dumay-Odelot H
    4. Da Silva D
    5. Rey C
    6. Prochazkova M
    7. Roeder RG
    8. Besser D
    9. Teichmann M

    (2010) Two isoforms of human RNA polymerase III with specific functions in cell growth and transformation

    PNAS 107:4176–4181.

    https://doi.org/10.1073/pnas.0914980107

    • PubMed
    • Google Scholar
22. 1. Heberle H
    2. Meirelles GV
    3. da Silva FR
    4. Telles GP
    5. Minghim R

    (2015) InteractiVenn: A web-based tool for the analysis of sets through Venn diagrams

    BMC Bioinformatics 16:169.

    https://doi.org/10.1186/s12859-015-0611-3

    • PubMed
    • Google Scholar
23. 1. Honjo RS
    2. Castro MAA
    3. Ferraciolli SF
    4. Soares Junior LAV
    5. Pastorino AC
    6. Bertola DR
    7. Miyake N
    8. Matsumoto N
    9. Kim CA

    (2021) Cerebellofaciodental syndrome in an adult patient: Expanding the phenotypic and natural history characteristics

    American Journal of Medical Genetics. Part A 185:1561–1568.

    https://doi.org/10.1002/ajmg.a.62140

    • PubMed
    • Google Scholar
24. 1. Jee YH
    2. Sowada N
    3. Markello TC
    4. Rezvani I
    5. Borck G
    6. Baron J

    (2017) BRF1 mutations in a family with growth failure, markedly delayed bone age, and central nervous system anomalies

    Clinical Genetics 91:739–747.

    https://doi.org/10.1111/cge.12887

    • PubMed
    • Google Scholar
25. 1. Johnson SS
    2. Zhang C
    3. Fromm J
    4. Willis IM
    5. Johnson DL

    (2007) Mammalian Maf1 is a negative regulator of transcription by all three nuclear RNA polymerases

    Molecular Cell 26:367–379.

    https://doi.org/10.1016/j.molcel.2007.03.021

    • PubMed
    • Google Scholar
26. 1. Kenneth NS
    2. Ramsbottom BA
    3. Gomez-Roman N
    4. Marshall L
    5. Cole PA
    6. White RJ

    (2007) TRRAP and GCN5 are used by c-Myc to activate RNA polymerase III transcription

    PNAS 104:14917–14922.

    https://doi.org/10.1073/pnas.0702909104

    • PubMed
    • Google Scholar
27. 1. Khanna A
    2. Johnson DL
    3. Curran SP

    (2014) Physiological roles for mafr-1 in reproduction and lipid homeostasis

    Cell Reports 9:2180–2191.

    https://doi.org/10.1016/j.celrep.2014.11.035

    • PubMed
    • Google Scholar
28. 1. Kutter C
    2. Brown GD
    3. Gonçalves A
    4. Wilson MD
    5. Watt S
    6. Brazma A
    7. White RJ
    8. Odom DT

    (2011) Pol III binding in six mammals shows conservation among amino acid isotypes despite divergence among tRNA genes

    Nature Genetics 43:948–955.

    https://doi.org/10.1038/ng.906

    • PubMed
    • Google Scholar
29. 1. Lata E
    2. Choquet K
    3. Sagliocco F
    4. Brais B
    5. Bernard G
    6. Teichmann M

    (2021) RNA Polymerase III Subunit Mutations in Genetic Diseases

    Frontiers in Molecular Biosciences 8:696438.

    https://doi.org/10.3389/fmolb.2021.696438

    • PubMed
    • Google Scholar
30. 1. Lee YL
    2. Li YC
    3. Su CH
    4. Chiao CH
    5. Lin IH
    6. Hsu MT

    (2015) MAF1 represses CDKN1A through a Pol III-dependent mechanism

    eLife 4:e06283.

    https://doi.org/10.7554/eLife.06283

    • PubMed
    • Google Scholar
31. 1. Levy R
    2. Levet C
    3. Cohen K
    4. Freeman M
    5. Mott R
    6. Iraqi F
    7. Gabet Y

    (2020) A genome-wide association study in mice reveals A role for Rhbdf2 in skeletal homeostasis

    Scientific Reports 10:3286.

    https://doi.org/10.1038/s41598-020-60146-8

    • PubMed
    • Google Scholar
32. 1. Li Y
    2. Tsang CK
    3. Wang S
    4. Li X-X
    5. Yang Y
    6. Fu L
    7. Huang W
    8. Li M
    9. Wang H-Y
    10. Zheng XFS

    (2016) MAF1 suppresses AKT-mTOR signaling and liver cancer through activation of PTEN transcription

    Hepatology (Baltimore, Md.) 63:1928–1942.

    https://doi.org/10.1002/hep.28507

    • PubMed
    • Google Scholar
33. 1. Lisignoli G
    2. Lambertini E
    3. Manferdini C
    4. Gabusi E
    5. Penolazzi L
    6. Paolella F
    7. Angelozzi M
    8. Casagranda V
    9. Piva R

    (2017) Collagen type XV and the “osteogenic status.”

    Journal of Cellular and Molecular Medicine 21:2236–2244.

    https://doi.org/10.1111/jcmm.13137

    • PubMed
    • Google Scholar
34. 1. Logan M
    2. Martin JF
    3. Nagy A
    4. Lobe C
    5. Olson EN
    6. Tabin CJ

    (2002) Expression of Cre Recombinase in the developing mouse limb bud driven by a Prxl enhancer

    Genesis (New York, N.Y 33:77–80.

    https://doi.org/10.1002/gene.10092

    • PubMed
    • Google Scholar
35. 1. McQueen C
    2. Hughes GL
    3. Pownall ME

    (2019) Skeletal muscle differentiation drives a dramatic downregulation of RNA polymerase III activity and differential expression of Polr3g isoforms

    Developmental Biology 454:74–84.

    https://doi.org/10.1016/j.ydbio.2019.06.001

    • PubMed
    • Google Scholar
36. 1. Meerbrey KL
    2. Hu G
    3. Kessler JD
    4. Roarty K
    5. Li MZ
    6. Fang JE
    7. Herschkowitz JI
    8. Burrows AE
    9. Ciccia A
    10. Sun T
    11. Schmitt EM
    12. Bernardi RJ
    13. Fu X
    14. Bland CS
    15. Cooper TA
    16. Schiff R
    17. Rosen JM
    18. Westbrook TF
    19. Elledge SJ

    (2011) The pINDUCER lentiviral toolkit for inducible RNA interference in vitro and in vivo

    PNAS 108:3665–3670.

    https://doi.org/10.1073/pnas.1019736108

    • PubMed
    • Google Scholar
37. 1. Mitra D
    2. Yasui OW
    3. Harvestine JN
    4. Link JM
    5. Hu JC
    6. Athanasiou KA
    7. Leach JK

    (2019) Exogenous Lysyl Oxidase-Like 2 and Perfusion Culture Induce Collagen Crosslink Formation in Osteogenic Grafts

    Biotechnology Journal 14:e1700763.

    https://doi.org/10.1002/biot.201700763

    • PubMed
    • Google Scholar
38. 1. Noack Watt KE
    2. Achilleos A
    3. Neben CL
    4. Merrill AE
    5. Trainor PA

    (2016) The Roles of RNA Polymerase I and III Subunits Polr1c and Polr1d in Craniofacial Development and in Zebrafish Models of Treacher Collins Syndrome

    PLOS Genetics 12:e1006187.

    https://doi.org/10.1371/journal.pgen.1006187

    • PubMed
    • Google Scholar
39. 1. Orioli A
    2. Pascali C
    3. Pagano A
    4. Teichmann M
    5. Dieci G

    (2012) RNA polymerase III transcription control elements: themes and variations

    Gene 493:185–194.

    https://doi.org/10.1016/j.gene.2011.06.015

    • PubMed
    • Google Scholar
40. 1. Orioli A
    2. Praz V
    3. Lhôte P
    4. Hernandez N

    (2016) Human MAF1 targets and represses active RNA polymerase III genes by preventing recruitment rather than inducing long-term transcriptional arrest

    Genome Research 26:624–635.

    https://doi.org/10.1101/gr.201400.115

    • PubMed
    • Google Scholar
41. 1. Palian BM
    2. Rohira AD
    3. Johnson SAS
    4. He L
    5. Zheng N
    6. Dubeau L
    7. Stiles BL
    8. Johnson DL

    (2014) Maf1 is a novel target of PTEN and PI3K signaling that negatively regulates oncogenesis and lipid metabolism

    PLOS Genetics 10:e1004789.

    https://doi.org/10.1371/journal.pgen.1004789

    • PubMed
    • Google Scholar
42. 1. Paolacci S
    2. Bertola D
    3. Franco J
    4. Mohammed S
    5. Tartaglia M
    6. Wollnik B
    7. Hennekam RC

    (2017) Wiedemann-Rautenstrauch syndrome: A phenotype analysis

    American Journal of Medical Genetics. Part A 173:1763–1772.

    https://doi.org/10.1002/ajmg.a.38246

    • PubMed
    • Google Scholar
43. 1. Pustylnik S
    2. Fiorino C
    3. Nabavi N
    4. Zappitelli T
    5. da Silva R
    6. Aubin JE
    7. Harrison RE

    (2013) EB1 levels are elevated in ascorbic Acid (AA)-stimulated osteoblasts and mediate cell-cell adhesion-induced osteoblast differentiation

    The Journal of Biological Chemistry 288:22096–22110.

    https://doi.org/10.1074/jbc.M113.481515

    • PubMed
    • Google Scholar
44. 1. Rak R
    2. Dahan O
    3. Pilpel Y

    (2018) Repertoires of tRNAs: The Couplers of Genomics and Proteomics

    Annual Review of Cell and Developmental Biology 34:239–264.

    https://doi.org/10.1146/annurev-cellbio-100617-062754

    • PubMed
    • Google Scholar
45. 1. Rowe PSN

    (2012) Regulation of bone-renal mineral and energy metabolism: the PHEX, FGF23, DMP1, MEPE ASARM pathway

    Critical Reviews in Eukaryotic Gene Expression 22:61–86.

    https://doi.org/10.1615/critreveukargeneexpr.v22.i1.50

    • PubMed
    • Google Scholar
46. 1. Saharia A
    2. Guittat L
    3. Crocker S
    4. Lim A
    5. Steffen M
    6. Kulkarni S
    7. Stewart SA

    (2008) Flap endonuclease 1 contributes to telomere stability

    Current Biology 18:496–500.

    https://doi.org/10.1016/j.cub.2008.02.071

    • PubMed
    • Google Scholar
47. 1. Schimmel P

    (2018) The emerging complexity of the tRNA world: mammalian tRNAs beyond protein synthesis

    Nature Reviews. Molecular Cell Biology 19:45–58.

    https://doi.org/10.1038/nrm.2017.77

    • PubMed
    • Google Scholar
48. 1. Shetty M
    2. Noguchi C
    3. Wilson S
    4. Martinez E
    5. Shiozaki K
    6. Sell C
    7. Mell JC
    8. Noguchi E

    (2020) Maf1-dependent transcriptional regulation of tRNAs prevents genomic instability and is associated with extended lifespan

    Aging Cell 19:1–13.

    https://doi.org/10.1111/acel.13068

    • PubMed
    • Google Scholar
49. 1. Sriskanthadevan-Pirahas S
    2. Deshpande R
    3. Lee B
    4. Grewal SS

    (2018) Ras/ERK-signalling promotes tRNA synthesis and growth via the RNA polymerase III repressor Maf1 in Drosophila

    PLOS Genetics 14:e1007202.

    https://doi.org/10.1371/journal.pgen.1007202

    • PubMed
    • Google Scholar
50. 1. Su Z
    2. Wilson B
    3. Kumar P
    4. Dutta A

    (2020) Noncanonical Roles of tRNAs: TRNA Fragments and Beyond

    Annual Review of Genetics 54:47–69.

    https://doi.org/10.1146/annurev-genet-022620-101840

    • PubMed
    • Google Scholar
51. 1. Sutcliffe JE
    2. Brown TRP
    3. Allison SJ
    4. Scott PH
    5. White RJ

    (2000) Retinoblastoma protein disrupts interactions required for RNA polymerase III transcription

    Molecular and Cellular Biology 20:9192–9202.

    https://doi.org/10.1128/MCB.20.24.9192-9202.2000

    • PubMed
    • Google Scholar
52. 1. Terhal PA
    2. Vlaar JM
    3. Middelkamp S
    4. Nievelstein RAJ
    5. Nikkels PGJ
    6. Ross J
    7. Créton M
    8. Bos JW
    9. Voskuil-Kerkhof ESM
    10. Cuppen E
    11. Knoers N
    12. van Gassen KLI

    (2020) Biallelic variants in POLR3GL cause endosteal hyperostosis and oligodontia

    European Journal of Human Genetics 28:31–39.

    https://doi.org/10.1038/s41431-019-0427-0

    • PubMed
    • Google Scholar
53. 1. Thomas A
    2. Thomas AK

    (2019) POLR3-related Leukodystrophy

    Journal of Clinical Imaging Science 9:45.

    https://doi.org/10.25259/JCIS_116_2019

    • Google Scholar
54. 1. Trainor PA
    2. Merrill AE

    (2014) Ribosome biogenesis in skeletal development and the pathogenesis of skeletal disorders

    Biochimica et Biophysica Acta 1842:769–778.

    https://doi.org/10.1016/j.bbadis.2013.11.010

    • PubMed
    • Google Scholar
55. 1. Valenzuela I
    2. Codina M
    3. Fernández-Álvarez P
    4. Mur P
    5. Valle L
    6. Tizzano EF
    7. Cuscó I

    (2020) Expanding the phenotype of cerebellar-facial-dental syndrome: Two siblings with a novel variant in BRF1

    American Journal of Medical Genetics. Part A 182:2742–2745.

    https://doi.org/10.1002/ajmg.a.61839

    • PubMed
    • Google Scholar
56. 1. Van Itallie CM
    2. Rogan S
    3. Yu A
    4. Vidal LS
    5. Holmes J
    6. Anderson JM

    (2006) Two splice variants of claudin-10 in the kidney create paracellular pores with different ion selectivities

    American Journal of Physiology. Renal Physiology 291:F1288–F1299.

    https://doi.org/10.1152/ajprenal.00138.2006

    • PubMed
    • Google Scholar
57. 1. Vannini A
    2. Ringel R
    3. Kusser AG
    4. Berninghausen O
    5. Kassavetis GA
    6. Cramer P

    (2010) Molecular basis of RNA polymerase III transcription repression by Maf1

    Cell 143:59–70.

    https://doi.org/10.1016/j.cell.2010.09.002

    • PubMed
    • Google Scholar
58. Preprint

    1. Vorländer MK
    2. Baudin F
    3. Moir RD
    4. Wetzel R
    5. Hagen WJH
    6. Willis IM
    7. Müller CW

    (2019) Structural Basis of RNA Polymerase III Transcription Repression by Maf1

    bioRxiv.

    https://doi.org/10.1101/859132

    • Google Scholar
59. 1. Walker-Kopp N
    2. Jackobel AJ
    3. Pannafino GN
    4. Morocho PA
    5. Xu X
    6. Knutson BA

    (2017) Treacher Collins syndrome mutations in Saccharomyces cerevisiae destabilize RNA polymerase I and III complex integrity

    Human Molecular Genetics 26:4290–4300.

    https://doi.org/10.1093/hmg/ddx317

    • PubMed
    • Google Scholar
60. 1. Wambach JA
    2. Wegner DJ
    3. Patni N
    4. Kircher M
    5. Willing MC
    6. Baldridge D
    7. Xing C
    8. Agarwal AK
    9. Vergano SAS
    10. Patel C
    11. Grange DK
    12. Kenney A
    13. Najaf T
    14. Nickerson DA
    15. Bamshad MJ
    16. Cole FS
    17. Garg A

    (2018) Bi-allelic POLR3A Loss-of-Function Variants Cause Autosomal-Recessive Wiedemann-Rautenstrauch Syndrome

    American Journal of Human Genetics 103:968–975.

    https://doi.org/10.1016/j.ajhg.2018.10.010

    • PubMed
    • Google Scholar
61. 1. Wang X
    2. Gerber A
    3. Chen WY
    4. Roeder RG

    (2020) Functions of paralogous RNA polymerase III subunits POLR3G and POLR3GL in mouse development

    PNAS 117:15702–15711.

    https://doi.org/10.1073/pnas.1922821117

    • PubMed
    • Google Scholar
62. 1. Wei Y
    2. Xu J
    3. Zhang W
    4. Wen Z
    5. Liu F

    (2016) RNA polymerase III component Rpc9 regulates hematopoietic stem and progenitor cell maintenance in zebrafish

    Development (Cambridge, England) 143:2103–2110.

    https://doi.org/10.1242/dev.126797

    • PubMed
    • Google Scholar
63. 1. White RJ
    2. Trouche D
    3. Martin K
    4. Jackson SP
    5. Kouzarides T

    (1996) Repression of RNA polymerase III transcription by the retinoblastoma protein

    Nature 382:88–90.

    https://doi.org/10.1038/382088a0

    • PubMed
    • Google Scholar
64. 1. Willis IM
    2. Moir RD
    3. Hernandez N

    (2018) Metabolic programming a lean phenotype by deregulation of RNA polymerase III

    PNAS 115:12182–12187.

    https://doi.org/10.1073/pnas.1815590115

    • PubMed
    • Google Scholar
65. 1. Woiwode A
    2. Johnson SAS
    3. Zhong S
    4. Zhang C
    5. Roeder RG
    6. Teichmann M
    7. Johnson DL

    (2008) PTEN represses RNA polymerase III-dependent transcription by targeting the TFIIIB complex

    Molecular and Cellular Biology 28:4204–4214.

    https://doi.org/10.1128/MCB.01912-07

    • PubMed
    • Google Scholar
66. 1. Wong RC-B
    2. Pollan S
    3. Fong H
    4. Ibrahim A
    5. Smith EL
    6. Ho M
    7. Laslett AL
    8. Donovan PJ

    (2011) A novel role for an RNA polymerase III subunit POLR3G in regulating pluripotency in human embryonic stem cells

    Stem Cells (Dayton, Ohio) 29:1517–1527.

    https://doi.org/10.1002/stem.714

    • PubMed
    • Google Scholar
67. 1. Wu L
    2. Pan J
    3. Thoroddsen V
    4. Wysong DR
    5. Blackman RK
    6. Bulawa CE
    7. Gould AE
    8. Ocain TD
    9. Dick LR
    10. Errada P
    11. Dorr PK
    12. Parkinson T
    13. Wood T
    14. Kornitzer D
    15. Weissman Z
    16. Willis IM
    17. McGovern K

    (2003) Novel small-molecule inhibitors of RNA polymerase III

    Eukaryotic Cell 2:256–264.

    https://doi.org/10.1128/EC.2.2.256-264.2003

    • PubMed
    • Google Scholar
68. 1. Wu T
    2. Hu E
    3. Xu S
    4. Chen M
    5. Guo P
    6. Dai Z
    7. Feng T
    8. Zhou L
    9. Tang W
    10. Zhan L
    11. Fu X
    12. Liu S
    13. Bo X
    14. Yu G

    (2021a) clusterProfiler 4.0: A universal enrichment tool for interpreting omics data

    Innovation (New York, N.Y.) 2:100141.

    https://doi.org/10.1016/j.xinn.2021.100141

    • PubMed
    • Google Scholar
69. 1. Wu SW
    2. Li L
    3. Feng F
    4. Wang L
    5. Kong YY
    6. Liu XW
    7. Yin C

    (2021b) Whole-exome sequencing reveals POLR3B variants associated with progeria-related Wiedemann-Rautenstrauch syndrome

    Italian Journal of Pediatrics 47:1–6.

    https://doi.org/10.1186/s13052-021-01112-6

    • Google Scholar
70. 1. Yeganeh M
    2. Hernandez N

    (2020) RNA polymerase III transcription as a disease factor

    Genes & Development 34:865–882.

    https://doi.org/10.1101/gad.333989.119

    • PubMed
    • Google Scholar

Author details

1. Ellen Phillips

   Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, United States

   Contribution

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

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0003-3654-2066

2. Naseer Ahmad

   Departments of Medicine and Pharmacological Sciences and Center for translational Medicine and Pharmacology, Icahn School of Medicine at Mount Sinai, New York, United States

   Contribution

   Data curation, Investigation, Methodology

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-9514-2703

3. Li Sun

   Departments of Medicine and Pharmacological Sciences and Center for translational Medicine and Pharmacology, Icahn School of Medicine at Mount Sinai, New York, United States

   Contribution

   Data curation, Investigation, Methodology

   Competing interests

   No competing interests declared

4. James Iben

   Molecular Genomics Core, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States

   Contribution

   Formal analysis, Software, Validation, Writing – review and editing

   Competing interests

   No competing interests declared

5. Christopher J Walkey

   Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, United States

   Contribution

   Conceptualization, Investigation, Resources

   Competing interests

   No competing interests declared

6. Aleksandra Rusin

   Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, United States

   Contribution

   Methodology, Supervision

   Competing interests

   No competing interests declared

7. Tony Yuen

   Departments of Medicine and Pharmacological Sciences and Center for translational Medicine and Pharmacology, Icahn School of Medicine at Mount Sinai, New York, United States

   Contribution

   Validation, Writing – review and editing

   Competing interests

   No competing interests declared

8. Clifford J Rosen

   Center for Clinical and Translational Research, Maine Medical Center Research Institute, Scarborough, United States

   Contribution

   Data curation, Formal analysis, Investigation

   Competing interests

   No competing interests declared

9. Ian M Willis

   Departments of Biochemistry and Systems and Computational Biology, Albert Einstein College of Medicine, Bronx, United States

   Contribution

   Project administration, Resources, Supervision, Writing – review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-6599-2395

10. Mone Zaidi

    Departments of Medicine and Pharmacological Sciences and Center for translational Medicine and Pharmacology, Icahn School of Medicine at Mount Sinai, New York, United States

    Contribution

    Investigation, Methodology, Supervision, Writing – review and editing

    Competing interests

    Senior editor, eLife

    "This ORCID iD identifies the author of this article:" 0000-0001-5911-9522

11. Deborah L Johnson

    Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, United States

    Contribution

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

    For correspondence

    deborah.johnson@bcm.edu

    Competing interests

    No competing interests declared

    "This ORCID iD identifies the author of this article:" 0000-0003-1745-1580

• 937

  views

• 286

  downloads

• 6

  citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.