Anti-TMUB1 monoclonal antibody Made to order

Misfolded proteins in the endoplasmic reticulum (ER) are degraded by ER-associated degradation (ERAD). Although ERAD components involved in degradation of luminal substrates are well characterized, much less is known about quality control of membrane proteins. Here, we analyzed the degradation pathways of two short-lived ER membrane model proteins in mammalian cells. Using a CRISPR-Cas9 genome-wide library screen, we identified an ERAD branch required for quality control of a subset of membrane proteins. Using biochemical and mass spectrometry approaches, we showed that this ERAD branch is defined by an ER membrane complex consisting of the ubiquitin ligase RNF185, the ubiquitin-like domain containing proteins TMUB1/2 and TMEM259/Membralin, a poorly characterized protein. This complex cooperates with cytosolic ubiquitin ligase UBE3C and p97 ATPase in degrading their membrane substrates. Our data reveal that ERAD branches have remarkable specificity for their membrane substrates, suggesting that multiple, perhaps combinatorial, determinants are involved in substrate selection.

Background Transmembrane and ubiquitin-like domain-containing 1 protein (Tmub1) negatively regulates liver regeneration. However, whether this regulation involves posttranscriptional modification of Tmub1 expression is unknown. Aim The aim of the study was to investigate whether microRNA (miR)-27a/b regulates posttranscriptional modification of Tmub1 and cell proliferation during liver regeneration. Methods Tmub1 mRNA 3'-untranslated region (UTR) sequences were analyzed using online software. A luciferase assay was used to verify the relationship between miR-27a/b and the 3'-UTR of Tmub1. Rat partial hepatectomy models were used to investigate miR-27a/b and Tmub1 levels after partial hepatectomy. MiR-27a/b expression was down- and up-regulated with mimics and inhibitors, respectively, to observe the effects of miR-27a/b on Tmub1 expression. Quantitative RT-PCR and Western blot analyses were used to measure miR-27a/b and Tmub1 expression. Hepatocyte proliferation was measured using the CCK8 method for BRL-3A liver cells and proliferating cell nuclear antigen and histone H3 phosphorylation in the regenerating liver. Results A potential binding site of miR-27a/b was found in the 3'-UTR sequence of Tmub1. Our luciferase assay confirmed that the Tmub1 mRNA 3'-UTR was the target of miR-27a/b. We observed a temporal correlation between miR-27a/b and Tmub1 expression during liver regeneration. MiR-27a/b down-regulated Tmub1 expression both in vivo and in vitro. MiR-27a/b regulated hepatocyte proliferation during liver regeneration. Conclusion MiR-27a/b regulates hepatocyte proliferation by controlling posttranscriptional modification of Tmub1 during liver regeneration.