Expression of nNOS alpha and beta isoforms in skeletal muscle of mice

Biochem J. 2024 Apr 9:BCJ20230458. doi: 10.1042/BCJ20230458. Online ahead of print.ABSTRACTKnowledge of the primary structure of neuronal NO synthase (nNOS) in skeletal muscle is still conflicting and needs further clarification. To elucidate the expression patterns of nNOS isoforms at both mRNA and protein level, systematic RT-PCR and epitope mapping by qualitative immunoblot analysis on skeletal muscle of C57/BL6 mice were performed. The ability of the nNOS isoforms to form aggregates was characterized by native low-temperature polyacrylamide electrophoresis (LT-PAGE). The molecular analysis was focused on the rectus femoris (RF) muscle, a skeletal muscle with a nearly balanced ratio of nNOS alpha- and beta-isoforms. RT-PCR amplificates from RF muscles showed exclusive exon-1d mRNA expression, either with or without exon-mu. Epitope mapping demonstrated the simultaneous expression of the nNOS splice variants alpha/mu, alpha/non-mu, beta/mu and beta/non-mu. Furthermore, immunoblotting suggests that the transition between nNOS alpha- and beta-isoforms lies within exon-3. In LT-PAGE, three protein nNOS associated aggregates iwere detected in homogenates of RF muscle and tibialis anterior muscle: a 320 kDa band containing nNOS alpha-isoforms, while 250 kDa and 300 kDa bands consist of nNOS beta-isoforms that form homodimers or heterodimers with non-nNOS proteins.PMID:38592741 | DOI:10.1042/BCJ20230458
Source: The Biochemical Journal - Category: Biochemistry Authors: Source Type: research