Partial molecular cloning, characterization, and analysis of the subcellular localization and expression patterns of the porcine OTUB1 gene

Abstract

The product of the OTUB1 gene is a member of the OTU superfamily of predicted cysteine proteases and inhibits cytokine gene transcription via its interaction with a ubiquitin protease and E3 ubiquitin ligase. To further understand the functions of the porcine OTUB1 gene, the subcellular localization of porcine OTUB1 protein was analyzed. We first cloned a partial DNA sequence of porcine OTUB1 which contained an 816 bp ORF encoding 271 amino acids. The deduced protein product was found to contain an OTU domain. The corresponding porcine OTUB1 protein was subsequently demonstrated to localize predominantly in the nucleus by confocal fluorescence microscopy. By spatial expression analysis, we further found that OTUB1 is highly expressed in the brain, liver, spleen, lung, kidney, large intestine, small intestine, stomach, ovary, uterus and thymus. In contrast, only low levels of this gene were evident in the heart, dorsal muscles and leg muscle of the pig. This is the first report to show the subcellular localization of porcine OTUB1, and our current data provides us with an important basis for conducting further studies on the functions and regulatory mechanisms underlying the role of OTUB1 gene in the immune system.

Structural basis and specificity of human otubain 1-mediated deubiquitination

“…We noted a considerable amount of co-immunoprecipitating nuclear proteins, such as cell growth nucleolar proteins, DNA topoisomerases and several RNA helicases sharing a DEAD (Asp-Glu-Ala-Asp) box motif ( Figure 6 and Supplementary Table S1). OTUB1 does contain a predicted NLS (nuclear localization sequence) (residues 69-85), and initial microscopy studies suggest that OTUB1 subsets are present in the nucleus ( [41] and A. Iphöfer, M. J. Edelmann and B. M. Kessler, unpublished work). On the other hand, RNA-binding proteins are abundant and commonly found as contaminants in MS-based pull-down assays [22], although all proteins listed were not detectable in controls without the bait protein ( Figure 6).…”

Differential Protein Expression in the Nucleus Accumbens and Amygdala of Lewis and Fischer 344 Rats, and its Relevance in Drug Addiction

“…OTUB1 also inhibits cytokine gene transcription through its interaction with a ubiquitin protease and E3 ubiquitin ligase [34]. After a careful search in the literature, we failed to find any direct link between this protein and drug addiction or any other relevant behavioral process.…”

Characterization of porcine SKIP gene in skeletal muscle development: Polymorphisms, association analysis, expression and regulation of cell growth in C2C12 cells

“…RNA extraction, cDNA synthesis, and real-time PCR Total RNA was extracted from the freshly harvested tissues and cells using a TRIZOL reagent kit (Invitrogen) according to the manufacturer's instructions. Reverse transcription was performed as described previously (Liu & Xiong, 2009;Shan et al, 2009). The PCR primer sequences used for real-time PCR for SKIP (GQ504265), β-actin (DQ845171.1), PCNA (NM_011045.2), p27KIP1 (NM_009875.4) and 18 s (NR_003278) are presented in Supplemental Table 1.…”