Porcine ubiquitin-like protein MNSFβ promotes cell apoptosis and covalently binds to BCL-G to enhance staurosporine-induced apoptosis

Abstract

Background: Monoclonal non-specific suppressor factor β (MNSFβ) is a ubiquitously expressed member of the ubiquitin-like family. It functions as a regulator of cell apoptosis and a potential tumor suppressor, playing a vital role in the processes of immune cell function and apoptosis.Methods: The present study constructed GFP-pMNSFβ swine umbilical vein endothelial cell (SUVEC) lines and investigated the function of porcine MNSFβ (pMNSFβ) in apoptosis, as well as its interactions with pBCL-G. Results revealed that stably expressed pMNSFβ protein in SUVEC lines significantly enhanced staurosporine (STS)-induced apoptosis. pMNSFβ proteins interacted with pBCL-G proteins and the expression of these interacting proteins synergized to further enhance STS-induced apoptosis.Results: GFP-pMNSFβ stably expressed SUVEC lines through transient transfection and neomycin screening methods. Over 90% of the SUVEC cultures expressed GFP signals, and 41.5 kDa GFP-pMNSFβ proteins were detected with western blotting methods. Annexin V-PE/PI staining and flow cytometry analyses showed that overexpression of pMNSFβ proteins significantly elevated STS-induced apoptosis rates. Co-immunoprecipitation methods revealed an interaction between pMNSFβ and pBCL-G proteins. BCL-G is a proapoptotic member of the BCL-2 family that has been shown to be misexpressed in human systemic lupus erythematosus, as well as mammary and prostate cancers. Here, we demonstrated that the co-expression and potential conjugation of pMNSFβ and pBCL-G proteins synergistically enhanced STSinduced apoptosis.Conclusions: The present study was the first to characterize the function of MNSFβ in porcine cells, and to clarify the function of MNSFβ in apoptosis. These results reveal that pMNSFβ is a potential molecular model for future investigations of diseases related to human MNSFβ dysfunction.

BCL-G: 20 years of research on a non-typical protein from the BCL-2 family

“…FUBI (Ubi-L) was shown to covalently bind to Bcl-G through an isopeptide bond between the C-terminal Gly 74 residue in FUBI (Ubi-L) and Lys 110 in Bcl-G [ 27 ]. Bcl-G-Ubi-L adduct was detected in the spleen, thymus, and brain, but not in the testes [ 27 ], indicating that its presence did not overlap with a tissue-specific abundance of Bcl-G. A similar association was demonstrated for porcine Mnsfβ and Bcl-G [ 53 ]. The significance of BCL-G modification by FUBI is unlikely to be associated with the formation of polyubiquitin-like chains to promote proteasomal degradation as Lys 48 is not conserved in ubiquitin-like proteins [ 52 ].…”