A novel approach to assess the ubiquitin‐fold modifier 1‐system in cells

Ishimura, Ryosuke; Obata, Miki; Kageyama, Shun; Daniel, Jens; Tanaka, Keiji; Komatsu, Masaaki

doi:10.1002/1873-3468.12518

Cited by 31 publications

(38 citation statements)

References 22 publications

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“…DDRGK1, the membrane anchor for the UFM1 E3 ligase complex, previously identified only following overexpression (5), was not identified in our UFMylome. Like the E3s for ubiquitin and other UBLs, DDRGK1 is subject to automodification by UFM1 when overexpressed (5,16). ASC1, a nuclear receptor coactivator reported to be a covalent UFMylation target (7), was also absent from our replicate datasets and has not been independently validated.…”

Section: Discussionmentioning

confidence: 97%

“…The use of the C-terminally truncated UFM1 bypasses the need to remove the C-terminal SerCys dipeptide from pro-UFM1 to expose the reactive C-terminal Gly (Fig. 1A) (6,16). Immunoblot analysis conducted under reducing and nonreducing conditions confirmed that 6xHis-UFM1 ΔSC formed thioester conjugates with the UFM1-specific E2, UFC1, in otherwise wildtype (UFM1 KO ;6xHis-UFM1 ΔSC ) and in double knockout (UFM1 KO , UFSP2 KO ) cells expressing 6xHis-UFM1 ΔSC , but not in double knockout (UFM1 KO , UBA5 KO ) cells expressing 6xHis-UFM1 ΔSC (SI Appendix, Fig.…”

Section: Significancementioning

confidence: 99%

“…In humans, hypomorphic loss-offunction alleles of genes that code for UFM1, UBA5, or UFPS2 are linked to diverse pathologies, including leukodystrophy (3), epileptic encephalopathy (11), spinocerebellar ataxia (12), and skeletal abnormalities (13)(14)(15), revealing an essential role for UFMylation in multiple organ systems. Although several cellular proteins have been reported to be UFMylation targets (5,7,(16)(17)(18), none of these have been demonstrated to form covalent adducts with UFM1 under physiological conditions and none have been plausibly linked to the cellular or organismal pathophysiologies associated with loss-of-function UFMylation mutants in humans or experimental animals.…”

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confidence: 99%

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Ribosomal protein RPL26 is the principal target of UFMylation

Walczak

Leto

Zhang

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

144

254

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Ubiquitin fold modifier 1 (UFM1) is a small, metazoan-specific, ubiquitin-like protein modifier that is essential for embryonic development. Although loss-of-function mutations in UFM1 conjugation are linked to endoplasmic reticulum (ER) stress, neither the biological function nor the relevant cellular targets of this protein modifier are known. Here, we show that a largely uncharacterized ribosomal protein, RPL26, is the principal target of UFM1 conjugation. RPL26 UFMylation and de-UFMylation is catalyzed by enzyme complexes tethered to the cytoplasmic surface of the ER and UFMylated RPL26 is highly enriched on ER membrane-bound ribosomes and polysomes. Biochemical analysis and structural modeling establish that UFMylated RPL26 and the UFMylation machinery are in close proximity to the SEC61 translocon, suggesting that this modification plays a direct role in cotranslational protein translocation into the ER. These data suggest that UFMylation is a ribosomal modification specialized to facilitate metazoan-specific protein biogenesis at the ER.

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Section: Discussionmentioning

confidence: 97%

Section: Significancementioning

confidence: 99%

mentioning

confidence: 99%

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Ribosomal protein RPL26 is the principal target of UFMylation

Walczak

Leto

Zhang

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

144

254

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“…Cell culture: UBA5 -knockout HEK293T cells were used in this study 7 23. Briefly, to generate UBA5 -knockout cells, UBA5 gRNA designed using the CRISPR Design tool (http://crispr.mit.edu/) was subcloned into pX330-U6-Chimeric_BB-CBh-hSpCas9 (Addgene #42230), a human codon-optimised SpCas9 and chimeric gRNA expression plasmid.…”

Section: Methodsmentioning

confidence: 99%

A homozygousUBA5pathogenic variant causes a fatal congenital neuropathy

et al. 2020

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BackgroundUBA5 is the activating enzyme of UFM1 in the ufmylation post-translational modification system. Different neurological phenotypes have been associated with UBA5 pathogenic variants including epilepsy, intellectual disability, movement disorders and ataxia.Methods and resultsWe describe a large multigenerational consanguineous family presenting with a severe congenital neuropathy causing early death in infancy. Whole exome sequencing and linkage analysis identified a novel homozygous UBA5 NM_024818.3 c.31C>T (p.Arg11Trp) mutation. Protein expression assays in mouse tissue showed similar levels of UBA5 in peripheral nerves to the central nervous system. CRISPR-Cas9 edited HEK (human embrionic kidney) cells homozygous for the UBA5 p.Arg11Trp mutation showed reduced levels of UBA5 protein compared with the wild-type. The mutant p.Arg11Trp UBA5 protein shows reduced ability to activate UFM1.ConclusionThis report expands the phenotypical spectrum of UBA5 mutations to include fatal peripheral neuropathy.

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“…Conversely, a search for inhibitors of Ufm1 proteases may identify a compound capable to enriching for ufmylated proteins. A recent report shows that loss of Ufsp2 in cells leads to a dramatic increase in ufmylated targets 16 , suggesting that inhibitors may yield the desired effect. Since heart failure is a leading cause of morbidity and mortality, especially among middle-aged and older adults, this important study reveals ufmylation as a promising pathway to explore for therapeutic benefits in cardiomyopathies, and likely other ER stress-based illnesses.…”

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confidence: 99%

Putting the Stress on UFM1 (Ubiquitin-Fold Modifier 1)

Sutherland

Barrio

2018

Circ: Heart Failure

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Defects in protein homeostasis (also known as proteostasis) are intrinsically linked to age-related decline of cardiac function and likely contribute to cardiomyopathies. Protein-folding chaperones and protein quality control systems work overtime in the high stress cardiac environment, responding to wear and tear. Tight regulation of the pathway components is often controlled by post-translational modifications (PTMs). In addition to compact PTMs such as phosphorylation, bulkier PTMs involve ubiquitin (Ub) and other small ubiquitin-like (UbL) proteins, which are covalently conjugated to target proteins. These modifications can modify stability, localization, and function of the target protein. Ufm1 (ubiquitin-fold modifier 1) is a less-studied UbL, but modification by Ufm1 (ufmylation 1) is emerging as an important mediator of the endoplasmic reticulum (ER) stress response, which is activated in cardiomyocytes during heart failure. Focusing on Ufl1 (Ufm1-ligase 1), one of the enzymes that mediate ufmylation, the report from Huabo Su and colleagues 2 provides strong evidence that Ufl1 has a cardioprotective role and points to the ufmylation pathway as a potential target for pharmacological intervention for some cardiomyopathies.

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A novel approach to assess the ubiquitin‐fold modifier 1‐system in cells

Cited by 31 publications

References 22 publications

Ribosomal protein RPL26 is the principal target of UFMylation

Ribosomal protein RPL26 is the principal target of UFMylation

A homozygousUBA5pathogenic variant causes a fatal congenital neuropathy

Putting the Stress on UFM1 (Ubiquitin-Fold Modifier 1)

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