MURF2B, a Novel LC3-Binding Protein, Participates with MURF2A in the Switch between Autophagy and Ubiquitin Proteasome System during Differentiation of C2C12 Muscle Cells

Abstract

The ubiquitin proteasome system and macroautophagy are proteolytic pathways essential in the maintenance of cellular homeostasis during differentiation and remodelling of skeletal muscle. In both pathways, proteins to be degraded are tagged with polyubiquitin. In skeletal muscles, the MURF2 proteins display E3 ubiquitin ligase structure suggesting that they may covalently attach ubiquitin polypeptides to still unknown target proteins. So far only MURF2A isoforms were studied and shown to interact with p62/SQSTM1, a protein implicated in macroautophagic and ubiquitin proteasome system degradations. Here, we analyzed the MURF2B and MURF2A proteins and show that the ratio of the isoforms changes during differentiation of muscle C2C12 cells and that the shift of the isoforms expression follows the sequential activation of autophagic or proteasomal degradation. We also show that MURF2B has a functional domain needed for its interaction with LC3, a protein needed for autophagic vesicles formation. Using specific MURF2 RNAi cells we observed that MURF2A and MURF2B are both needed for the formation of autophagosomes and that in the absence of MURF2B, the cells expressing MURF2A display an activated ubiquitin proteasome system implicated in the degradation of p62/SQSTM1 by UPS. Altogether, our results indicate that MURF2A and MURF2B proteins could participate in the molecular switch between the two ubiquitin degradative pathways.

Precision autophagy directed by receptor regulators – emerging examples within the TRIM family

“…Other TRIMs interact with mAtg8s (Table S1) in vitro, as shown for TRIM16, TRIM17, TRIM22, TRIM49 and TRIM55 (Mandell et al, 2014). Colocalization between mAtg8s and the TRIMs listed above in the cytoplasm (Niida et al, 2010;Pizon et al, 2013;Mandell et al, 2014;Kimura et al, 2015), as well as with TRIM63 , corroborate the biochemical findings. Potential binding to mAtg8 has also been documented for TRIM19 [also known as promyelocytic leukemia (PML) protein] and TRIM28 (Behrends et al, 2010;He et al, 2014).…”

“…2C; Table S1) (Lipinski et al, 2010;Niida et al, 2010;Perera et al, 2011;McKnight et al, 2012;Tomar et al, 2012;Barde et al, 2013;Pizon et al, 2013;Yang et al, 2013;Khan et al, 2014;Mandell et al, 2014;Choi et al, 2015;Kimura et al, 2015). TRIMs act as autophagy receptor regulators (meaning that they act as both receptors and platforms for assembly of autophagosome machinery) by two means.…”

“…The TRIMs show a preference for GABARAPs and LC3A, and do not bind to LC3B when tested in vitro. Nevertheless, LC3B can be found in larger protein complexes with TRIMs when precipitated from cells (Pizon et al, 2013;He et al, 2014;Mandell et al, 2014). Thus, it is likely that LC3B forms larger complexes with TRIMs, possibly indirectly and in a manner that is mediated by an intermediary protein (e.g.…”

“…3A); the known examples include TRIM5 (O'Connor et al, 2010), TRIM50 , TRIM55 (Lange et al, 2005;Pizon et al, 2013;Mandell et al, 2014), TRIM13 (Tomar et al, 2012), TRIM17 and TRIM49 (Mandell et al, 2014), TRIM21 , as well as TRIM76 (Blandin et al, 2013). Furthermore, p62 can colocalize with TRIM63 in the cytoplasm and with TRIM19 in the nucleus within the nuclear PML bodies (Pankiv et al, 2010), although this might not be restricted to the nucleus because TRIM19 functions in the cytoplasm as well (Lin et al, 2004;Dutrieux et al, 2015).…”

“…TRIMs affect autophagy as a whole process (Lipinski et al, 2010;Perera et al, 2011;McKnight et al, 2012;Tomar et al, 2012;Barde et al, 2013;Pizon et al, 2013;Khan et al, 2014;Pineda et al, 2015) and interact with ATG factors (Behrends et al, 2010;Yang et al, 2013;Mandell et al, 2014;Kimura et al, 2015). Of the 82 human TRIMs, 49 TRIMs have an effect on autophagy (Table S1).…”