Limited column formation in the embryonic growth plate implies divergent growth mechanisms during pre- and postnatal bone development
- Weizmann Institute of Science, Israel
- University of Michigan-Ann Arbor, United States
- Aix Marseille Univ, INSERM, MMG, UMR1251, France
- Queen Mary University of London, United Kingdom
Abstract
Chondrocyte columns, which are a hallmark of growth plate architecture, play a central role in bone elongation. Columns are formed by clonal expansion following rotation of the division plane, resulting in a stack of cells oriented parallel to the growth direction. In this work, we analyzed hundreds of Confetti multicolor clones in growth plates of mouse embryos using a pipeline comprising 3D imaging and algorithms for morphometric analysis. Surprisingly, analysis of the elevation angles between neighboring pairs of cells revealed that most cells did not display the typical stacking pattern associated with column formation, implying incomplete rotation of the division plane. Morphological analysis revealed that although embryonic clones were elongated, they formed clusters oriented perpendicular to the growth direction. Analysis of growth plates of postnatal mice revealed both complex columns, composed of ordered and disordered cell stacks, and small, disorganized clusters located in the outer edges. Finally, correlation between the temporal dynamics of the ratios between clusters and columns and between bone elongation and expansion suggests that clusters may promote expansion, whereas columns support elongation. Overall, our findings support the idea that modulations of division plane rotation of proliferating chondrocytes determines the formation of either clusters or columns, a multifunctional design that regulates morphogenesis throughout pre- and postnatal bone growth. Broadly, this work provides a new understanding of the cellular mechanisms underlying growth plate activity and bone elongation during development.
Data availability
The datasets generated and analyzed during the current study are available on Zenodo at the following links: https://doi.org/10.5281/zenodo.10440013, https://doi.org/10.5281/zenodo.10444731, https://doi.org/10.5281/zenodo.10446055, https://doi.org/10.5281/zenodo.10446092, https://doi.org/10.5281/zenodo.10446121, https://doi.org/10.5281/zenodo.10446131, https://doi.org/10.5281/zenodo.10446123, https://doi.org/10.5281/zenodo.10446145.
-
Col2creER;Confettihet_gE14.5_E18.5_litter154_m3Zenodo, 10.5281/zenodo.10440013.
-
Col2creER;Confettihet_gE14.5_E18.5_litter154_m4enodo, 10.5281/zenodo.10444731.
-
Col2creER;Confettihet_gE14.5_E18.5_litter153_m7enodo, 10.5281/zenodo.10446055.
-
Col2creER;Confettihet_gP30_P40_litter152_m3_DFenodo, 10.5281/zenodo.10446092.
-
Col2creER;Confettihet_gP30_P40_litter152_m3_PTenodo, 10.5281/zenodo.10446121.
-
Col2creER;Confettihet_gP30_P40_litter152_m4_DFenodo, 10.5281/zenodo.10446131.
-
Col2creER;Confettihet_gP30_P40_litter151_m2enodo, 10.5281/zenodo.10446123.
-
Col2creER;Confettihet_gP30_P40_litter152_m4_PTenodo, 10.5281/zenodo.10446145.
Article and author information
Author details
Funding
Israel Science Foundation (1387/23)
- Elazar Zelzer
Weizmann - Sagol Institute for longetivity research
- Elazar Zelzer
Julie and Eric Borman Family Research Funds
- Elazar Zelzer
University of Michigan School of Dentistry startup funds
- Tomer Stern
University of Michigan Oral Health Sciences PhD program
- Meng-Jia Lian
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: All animal experiments were pre-approved by and conducted according to the guidelines of the Institutional Animal Care and Use Committee (IACUC) of the Weizmann Institute (IACUC 01750221-1 and IACUC 05700723-2). All animals used in this study had access to food and water ad libitum and were maintained under controlled humidity and temperature (45-65%, 22 {plus minus} 2{degree sign}C, respectively).
Copyright
© 2024, Rubin et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 1,187
- views
-
- 173
- downloads
-
- 0
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Limited column formation in the embryonic growth plate implies divergent growth mechanisms during pre- and postnatal bone development
eLife 13:e95289.
https://doi.org/10.7554/eLife.95289
Further reading
-
- Cell Biology
How the fate (folding versus degradation) of glycoproteins is determined in the endoplasmic reticulum (ER) is an intriguing question. Monoglucosylated glycoproteins are recognized by lectin chaperones to facilitate their folding, whereas glycoproteins exposing well-trimmed mannoses are subjected to glycoprotein ER-associated degradation (gpERAD); we have elucidated how mannoses are sequentially trimmed by EDEM family members (George et al., 2020; 2021 eLife). Although reglucosylation by UGGT was previously reported to have no effect on substrate degradation, here we directly tested this notion using cells with genetically disrupted UGGT1/2. Strikingly, the results showed that UGGT1 delayed the degradation of misfolded substrates and unstable glycoproteins including ATF6α. An experiment with a point mutant of UGGT1 indicated that the glucosylation activity of UGGT1 was required for the inhibition of early glycoprotein degradation. These and overexpression-based competition experiments suggested that the fate of glycoproteins is determined by a tug-of-war between structure formation by UGGT1 and degradation by EDEMs. We further demonstrated the physiological importance of UGGT1, since ATF6α cannot function properly without UGGT1. Thus, our work strongly suggests that UGGT1 is a central factor in ER protein quality control via the regulation of both glycoprotein folding and degradation.
-
- Biochemistry and Chemical Biology
- Cell Biology
Activation of the Wnt/β-catenin pathway crucially depends on the polymerization of dishevelled 2 (DVL2) into biomolecular condensates. However, given the low affinity of known DVL2 self-interaction sites and its low cellular concentration, it is unclear how polymers can form. Here, we detect oligomeric DVL2 complexes at endogenous protein levels in human cell lines, using a biochemical ultracentrifugation assay. We identify a low-complexity region (LCR4) in the C-terminus whose deletion and fusion decreased and increased the complexes, respectively. Notably, LCR4-induced complexes correlated with the formation of microscopically visible multimeric condensates. Adjacent to LCR4, we mapped a conserved domain (CD2) promoting condensates only. Molecularly, LCR4 and CD2 mediated DVL2 self-interaction via aggregating residues and phenylalanine stickers, respectively. Point mutations inactivating these interaction sites impaired Wnt pathway activation by DVL2. Our study discovers DVL2 complexes with functional importance for Wnt/β-catenin signaling. Moreover, we provide evidence that DVL2 condensates form in two steps by pre-oligomerization via high-affinity interaction sites, such as LCR4, and subsequent condensation via low-affinity interaction sites, such as CD2.
