Bit-1 is an essential regulator of myogenic differentiation

Abstract

Muscle differentiation requires a complex signaling cascade that leads to the production of multinucleated myofibers. Genes regulating the intrinsic mitochondrial apoptotic pathway also function in controlling cell differentiation. How such signaling pathways are regulated during differentiation is not fully understood. Bit-1 (also known as PTRH2) mutations in humans cause infantile-onset multisystem disease with muscle weakness. We demonstrate here that Bit-1 controls skeletal myogenesis through a caspase-mediated signaling pathway. Bit-1-null mice exhibit a myopathy with hypotrophic myofibers. Bit-1-null myoblasts prematurely express muscle-specific proteins. Similarly, knockdown of Bit-1 expression in C2C12 myoblasts promotes early differentiation, whereas overexpression delays differentiation. In wildtype mice, Bit-1 levels increase during differentiation. Bit-1-null myoblasts exhibited increased levels of caspase 9 and caspase 3 without increased apoptosis. Bit-1 re-expression partially rescued differentiation. In Bit-1-null muscle, Bcl-2 levels are reduced, suggesting that Bcl-2-mediated inhibition of caspase 9 and caspase 3 is decreased. Bcl-2 re-expression rescued Bit-1-mediated early differentiation in Bit-1-null myoblasts and C2C12 cells with knockdown of Bit-1 expression. These results support an unanticipated yet essential role for Bit-1 in controlling myogenesis through regulation of Bcl-2.

The beneficial role of proteolysis in skeletal muscle growth and stress adaptation

“…Here, Fernando et al [ 29 ] demonstrated that transient caspase-3 activity is required for myoblast differentiation and that this non-death activity is mediated in part through the cleavage activation of the Ste-20 like kinase, macrophage stimulating 1 (MST1). Subsequent studies have established that the key elements of the intrinsic mediated cell death pathway are fully conserved to engage caspase 3 during myoblast differentiation [ 37 , 77 ]. Once activated, caspase 3 targets multiple substrates to engage the differentiation program (Fig.…”

“…These observations confirm that caspase 3 acts at multiple points to secure the differentiation program, yet the mechanism that spurs caspase activity at these specific temporal junctures remains unknown. Satellite cell activation has been reported to trigger engagement of the extrinsic mediated cell death pathway such as the Fas-associated protein with death domain (FADD) receptor [ 14 ], yet the requisite initiator caspase for this pathway, caspase 8, does not appear to be appreciably activated at this stage [ 37 ]. Given the central role of caspase 3 signaling in the differentiation process, there is a significant need to identify (1) the pathways that engage the activation of this protease, (2) the full range of substrates that facilitate its capacity to induce differentiation, and (3) the mechanisms that restrain protease activity and direct it to a non-death cell function.…”

“…As noted above, caspase-mediated cell differentiation is a broadly conserved mechanism that spans all cell lineages and is extant from worms to humans [ 24 ]. The key distinction between death and non-death caspase function is the duration of the signaling cascade, where cell death is characterized by sustained caspase 3/7 activity, non-death responses by a transient activity pattern [ 23 , 37 , 77 ]. A probable loci of caspase control may reside with the inhibitor of apoptosis (IAPs), a class of RING Finger domain proteins that target and inhibit caspase activity by both structural blockade and through self directed ubiquitination of the IAP/caspase complex [ 21 , 63 , 107 ].…”

PTRH2: an adhesion regulated molecular switch at the nexus of life, death, and differentiation

“…Ptrh2-null myoblasts exhibit earlier and higher expression levels of troponin T, myosin-2, and myogenin compared to age-matched controls, suggesting premature differentiation occurs when PTRH2 protein is absent. Reexpression of PTRH2 in null myoblasts rescues early differentiation and decreases caspase activity similar to littermate age-matched controls 44 .…”

Homozygous mutation in PTRH2 gene causes progressive sensorineural deafness and peripheral neuropathy

“…Overall, it was concluded that the UBL‐UBA shuttling pathway in the ubiquitin‐proteasome pathway is negatively regulated by PTRH2 (Ishii, Funakoshi, & Kobayashi, ). Notably, in mammalian cells, two additional functions of PTRH2 were reported: (1) Promoting cell survival via activation of both the PI3K‐AKT‐NFkB pathway and of Bcl‐2 transcription (Griffiths et al, , ); and (2) Promoting cell death by induction of anoikis (cell death induced upon cell detachment) (Kairouz‐Wahbe et al, ), which requires release of the protein from mitochondria and formation of a complex with the transcriptional regulator amino‐terminal enhancer of split (AES). Thus, PTRH2 is a multifunctional protein that participates in various cellular functions in various locations in the cell (Jan et al, ).…”