title: Ca++-sensitizing mutations in troponin, P(i), and 2-deoxyATP alter the depressive effect of acidosis on regulated thin-filament velocity.
abstract: Repeated, intense contractile activity compromises the ability of skeletal muscle to generate force and velocity, resulting in fatigue. The decrease in velocity is thought to be due, in part, to the intracellular build-up of acidosis inhibiting the function of the contractile proteins myosin and troponin; however, the underlying molecular basis of this process remains poorly understood. We sought to gain novel insight into the decrease in velocity by determining whether the depressive effect of acidosis could be altered by 1) introducing Ca(++)-sensitizing mutations into troponin (Tn) or 2) by agents that directly affect myosin function, including inorganic phosphate (Pi) and 2-deoxy-ATP (dATP) in an in vitro motility assay. Acidosis reduced regulated thin-filament velocity (VRTF) at both maximal and submaximal Ca(++) levels in a pH-dependent manner. A truncated construct of the inhibitory subunit of Tn (TnI) and a Ca(++)-sensitizing mutation in the Ca(++)-binding subunit of Tn (TnC) increased VRTF at submaximal Ca(++) under acidic conditions but had no effect on VRTF at maximal Ca(++) levels. In contrast, both Pi and replacement of ATP with dATP reversed much of the acidosis-induced depression of VRTF at saturating Ca(++). Interestingly, despite producing similar magnitude increases in VRTF, the combined effects of Pi and dATP were additive, suggesting different underlying mechanisms of action. These findings suggest that acidosis depresses velocity by slowing the detachment rate from actin but also by possibly slowing the attachment rate.
authors.affiliation: Department of Kinesiology, University of Massachusetts, Amherst, Massachusetts;
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Longyear
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Turner
authors.forename: Jonathan P
authors.initials: JP
authors.lastname: Davis
authors.forename: Joseph
authors.initials: J
authors.lastname: Lopez
authors.forename: Brandon
authors.initials: B
authors.lastname: Biesiadecki
authors.forename: Edward P
authors.initials: EP
authors.lastname: Debold
mesh_terms: D000138:Acidosis / Q000235:genetics* / Q000378:metabolism; D000199:Actins / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002645:Chickens; D003838:Deoxyadenine Nucleotides / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D009218:Myosins / Q000737:chemistry / Q000235:genetics; D010710:Phosphates / Q000502:physiology*; D017433:Protein Structure, Secondary; D011817:Rabbits; D014336:Troponin / Q000737:chemistry / Q000235:genetics*
chemical_list: D000199:Actins; D003838:Deoxyadenine Nucleotides; D010710:Phosphates; D014336:Troponin; D009218:Myosins; C026600:2'-deoxyadenosine triphosphate; D002118:Calcium
keywords: 2-deoxy-ATP; fatigue; phosphate; regulation; troponin C; troponin I