Search

Metazoan development from a one-cell zygote to a fully formed organism requires complex cellular differentiation and communication pathways. To coordinate these processes, embryos frequently encode signaling information with the small protein modifier ubiquitin, which is typically attached to lysine residues within substrates. During ubiquitin signaling, a three-step enzymatic cascade modifies specific substrates with topologically unique ubiquitin modifications, which mediate changes in the substrate’s stability, activity, localization, or interacting proteins. Ubiquitin signaling is critically regulated by deubiquitylases (DUBs), a class of ~100 human enzymes that oppose the conjugation of ubiquitin. DUBs control many essential cellular functions and various aspects of human physiology and development. Recent genetic studies have identified mutations in several DUBs that cause developmental disorders. Here we review principles controlling DUB activity and substrate recruitment that allow these enzymes to regulate ubiquitin signaling during development. We summarize key mechanisms of how DUBs control embryonic and postnatal differentiation processes, highlight developmental disorders that are caused by mutations in particular DUB members, and describe our current understanding of how these mutations disrupt development. Finally, we discuss how emerging tools from human disease genetics will enable the identification and study of novel congenital disease-causing DUBs.

The nematode C. elegans has a contingent of five sod genes, one of the largest among aerobic organisms. Earlier studies revealed each of the five sod genes is capable of making perfectly active SOD proteins in heterologous expression systems therefore none appears to be a pseudogene. Yet deletion of the entire contingent of sod genes fails to impose any effect on the survival of C. elegans except these animals appear more sensitive to extraneously applied oxidative stress conditions. We asked how many of the five sod genes are actually making active SOD enzymes in C. elegans through the usage of in-gel SOD activity analysis and by using KCN as a selective inhibitor against Cu-ZnSOD enzyme(s). Here we provide evidence that out of the five SOD proteins only the mitochondrial SOD is active in the water-soluble fraction of C. elegans extracts albeit at an apparently much lower activity than the multiple active SODs in D. melanogaster and E. coli. We had no opportunity to test the activity of Sod-4a isoform which is possibly a membrane-bound form of SOD. The mutant analysis further confirmed that among the two mitochondrial SOD proteins, SOD-2 is the only naturally active SOD in C. elegans.

Disease-causing mutations in a linkage-specific deubiquitylase provide insights into chromatin remodeling during embryogenesis., Reversible modification of proteins with linkage-specific ubiquitin chains is critical for intracellular signaling. Information on physiological roles and underlying mechanisms of particular ubiquitin linkages during human development are limited. Here, relying on genomic constraint scores, we identify 10 patients with multiple congenital anomalies caused by hemizygous variants in OTUD5, encoding a K48/K63 linkage–specific deubiquitylase. By studying these mutations, we find that OTUD5 controls neuroectodermal differentiation through cleaving K48-linked ubiquitin chains to counteract degradation of select chromatin regulators (e.g., ARID1A/B, histone deacetylase 2, and HCF1), mutations of which underlie diseases that exhibit phenotypic overlap with OTUD5 patients. Loss of OTUD5 during differentiation leads to less accessible chromatin at neuroectodermal enhancers and aberrant gene expression. Our study describes a previously unidentified disorder we name LINKED (LINKage-specific deubiquitylation deficiency–induced Embryonic Defects) syndrome and reveals linkage-specific ubiquitin cleavage from chromatin remodelers as an essential signaling mode that coordinates chromatin remodeling during embryogenesis.

The nematodeC. eleganshas a contingent of fivesodgenes, one of the largest among aerobic organism. Earlier studies revealed each of the fivesodgenes is capable of making perfectly active SOD proteins in heterologous expressions systems therefore none appears to be a pseudogene. Yet deletion of the entire contingent ofsodgenes fails to impose any effect on the survival ofC. elegansexcept these animals appear more sensitive to extraneously applied oxidative stress condition. We asked how many of the fivesodgenes are actually active inC. elegansthrough an in-gel SOD activity analysis. Here we provide evidence that out of the five genes only the mitochondrial SOD gene is active inC. elegans, albeit at a much lesser amount compared toD. melanogasterandE. coli. Mutant analysis further confirmed that among the mitochondrial forms, SOD-2 is the only naturally active SOD inC. elegans.

Embryonic development occurs through commitment of pluripotent stem cells to differentiation programs that require highly coordinated changes in gene expression. Chromatin remodeling of gene regulatory elements is a critical component of how such changes are achieved. While many factors controlling chromatin dynamics are known, mechanisms of how different chromatin regulators are orchestrated during development are not well understood. Here, we describe LINKED (LINKage-specific-deubiquitylation-deficiency-induced Embryonic Defects) syndrome, a novel multiple congenital anomaly disorder caused by hypomorphic hemizygous missense variants in the deubiquitylase OTUD5/DUBA. Studying LINKED mutations in vitro, in mouse, and in models of neuroectodermal differentiation of human pluripotent stem cells, we uncover a novel regulatory circuit that coordinates chromatin remodeling pathways during early differentiation. We show that the K48-linkage-specific deubiquitylation activity of OTUD5 is essential for murine and human development and, if reduced, leads to aberrant cell-fate specification. OTUD5 controls differentiation through preventing the degradation of multiple chromatin regulators including ARID1A/B and HDAC2, mutation of which underlie developmental syndromes that exhibit phenotypic overlap with LINKED patients. Accordingly, loss of OTUD5 during early differentiation leads to less accessible chromatin at neural and neural crest enhancers and thus aberrant rewiring of gene expression networks. Our work identifies a novel mechanistic link between phenotypically related developmental disorders and an essential function for linkagespecific ubiquitin editing of substrate groups (i.e. chromatin remodeling complexes) during early cellfate decisions – a regulatory concept, we predict to be a general feature of embryonic development.

Dietary proteins are crucial for oogenesis. The Target of Rapamycin (TOR) is a major nutrient sensor controlling organismal growth and fertility, but the downstream effectors of TOR signaling remain largely uncharacterized. We previously identified Drosophila Spargel/dPGC-1 as a terminal effector of the TOR-TSC pathway, and now report that Spargel connects nutrition to oogenesis. We found that Spargel is expressed predominantly in the ovaries of adult flies, and germline spargel knockdown inhibits cyst growth, ultimately leading to egg chamber degeneration and female sterility. In situ staining demonstrated nuclear localization of Spargel in the nurse cells and follicle cells of the ovariole. Furthermore, Spargel/dPGC-1 expression is influenced by dietary yeast concentration and TOR signaling, suggesting Spargel/dPGC-1 might transmit nutrient-mediated signals into ovarian growth. We propose that potentiating Spargel/dPGC-1 expression in the ovary is instrumental in nutrient-mediated regulation of oogenesis.

Peroxisome Proliferator Activated Receptor Gamma Co-activator-1 (PGC-1) is a well-conserved protein among all chordates. Entire Drosophila species subgroup carries a PGC-1 homolog in their genome called spargel/dPGC-1 showing very little divergence. Recent studies have reported that significant functional similarities are shared between vertebrate and invertebrate PGC-1's based on their role in mitochondrial functions and biogenesis, gluconeogenesis, and most likely in transcription and RNA processing. With the help of genetic epistasis analysis, we established that Drosophila Spargel/dPGC-1 affects cell growth process as a terminal effector in the Insulin-TOR signaling pathway. The association between Spargel/dPGC-1 and Insulin signaling could also explain its role in the aging process. Here we provided a further comparison between Spargel/dPGC-1 and PGC-1 focusing on nuclear localization, oxidative stress resistance, and a possible role of Spargel/dPGC-1 in oogenesis reminiscing the role of Spargel in reproductive aging like many Insulin signaling partners. This led us to hypothesize that the discovery of newer biological functions in Drosophila Spargel/dPGC-1 will pave the way to uncover novel functional equivalents in mammals.

Background Thus far, a handful of genes have been shown to be related to the wing maturation process in insects. A novel heme peroxidase enzyme known as curly suppressor (Cysu)(formerly CG5873), have been characterized in this report because it is involved in wing morphogenesis. Using bioinformatics tools we found that Cysu is remarkably conserved in the genus Drosophila (>95%) as well as in invertebrates (>70%), although its vertebrate orthologs show poor homology. Time-lapse imaging and histochemical analyses have confirmed that the defective wing phenotype of Cysu is not a result of any underlying cellular alterations; instead, its wings fail to expand in mature adults. Results The precise requirement of Cysu in wings was established by identifying a bona fide mutant of Cysu from the Bloomington Drosophila Stock Centre collection. Its requirement in the wing has also been shown by RNA knockdown of the gene. Subsequent transgenic rescue of the mutant wing phenotype with the wild-type gene confirmed the phenotype resulting from Cysu mutant. With appropriate GAL4 driver like engrailed-GAL4, the Cysu phenotype was compartmentalized, which raises a strong possibility that Cysu is not localized in the extracellular matrix (ECM); hence, Cysu is not engaged in bonding the dorsal and ventral cuticular layers. Finally, shortened lifespan of the Cysu mutant suggests it is functionally essential for other biological processes as well. Conclusion Cysu, a peroxinectin-like gene, is required during the wing maturation process in Drosophila because as a heme peroxidase, Cysu is capable of utilizing H2O2, which plays an essential role in post-eclosion wing morphogenesis. Electronic supplementary material The online version of this article (doi:10.1186/s12861-016-0143-8) contains supplementary material, which is available to authorized users.