Modulation of skeletal muscle fiber growth and specialization by mitogen-activated protein kinase pathways

Hao Shi, Purdue University

Abstract

The overall objective of this research was to investigate the role of mitogen-activated protein kinase (MAPK) pathways in the modulation of skeletal muscle hypertrophy, atrophy and fiber type determination. MAPK signaling has been widely studied during myogenesis and exercise, yet its role in muscle growth and specialization remains poorly understood. In the first study, we established an adult muscle hypertrophy model by feeding 20 ppm clenbuterol in drinking water to mice for two weeks. Results showed β-adrenergic receptor agonist (BA)-induced hypertrophy is muscle type-specific, more profound in fast-twitch skeletal muscles. Analysis of the β-adrenergic receptor signaling and its downstream pathways in vitro and in vivo revealed that the extracellular signal-regulated kinase-1, and -2 (ERK1/2) was differentially activated in slow- and fast-twitch muscles. Overexpression of the ERK1/2 deactivator MAPK phosphatase-1 (MKP1) blunted BA-induced hypertrophy in vivo. These findings suggest that the differential involvement of the basal and BA-induced active ERK1/2 may account, at least in part, for the observed differential hypertrophy in slow and fast muscles. Another interesting finding was that BA induced a muscle fiber type shift to the fastest phenotype as evidenced by the de novo synthesis of fastest myosin heavy chain IIb isoform, even in the soleus. These findings led us to hypothesize that MAPK drives the formation of the fast fiber phenotype. In the second study, we used slow and fast fiber type-specific reporters in cultured C2C12 myoblast cultures treated with MAPK inhibitors or constitutively expressing active ERK2. Results demonstrated that ERK1/2 is necessary for the fast fiber phenotype and repressed the slow fiber program. We confirmed these results in vivo by overexpressing an EGFP-tagged MKP1 in the superficial gastrocnemius muscles. Forced expression of MKP1 induced de novo synthesis of MyHC IIa and I in IIb/x fibers. This study helped explain the molecular control of fiber type transition in our first study. Curiously, we observed that those IIb/x fibers both EGFPMKP1 and MyHC IIa/I positive were generally smaller and therefore designed a study to address this apparent reduction in muscle cell size. Results of these studies clearly demonstrated that inhibition of ERK1/2 signaling induced atrophic responses in cultured myocytes and this atrophic process was mediated by the activation of the ubiquitin-proteosome proteolytic pathway and downregulation of Akt and its downstream signaling cascades. Over-expression of MKP1 induced atrophy in both soleus and gastrocnemius muscles in a time-dependent manner. Analysis of NFkB and 26S proteosome activity reporters revealed that they are differentially involved in MAPK-inhibition-induced atrophy in slow and fast muscles. Taken together, MAPK signaling plays an important role in muscle growth and fiber type specialization.

Degree

Ph.D.

Advisors

Grant, Purdue University.

Subject Area

Molecular biology|Cellular biology

COinS