An alternative domain near the nucleotide-binding site of Drosophila muscle myosin affects ATPase kinetics.
In Drosophila melanogaster expression of muscle myosin heavy chain isoforms occurs by alternative splicing of transcripts from a single gene. The exon 7 domain is one of four variable regions in the catalytic head and is located near the nucleotide-binding site. To ascribe a functional role to this domain, we created two chimeric myosin isoforms (indirect flight isoform-exon 7a and embryonic-exon 7d) that differ from the native indirect flight muscle and embryonic body-wall muscle isoforms only in the exon 7 region. Germline transformation and subsequent expression of the chimeric myosins in the indirect flight muscle of myosin-null Drosophila allowed us to purify the myosin for in vitro studies and to assess in vivo structure and function of transgenic muscles. Intriguingly, in vitro experiments show the exon 7 domain modulates myosin ATPase activity but has no effect on actin filament velocity, a novel result compared to similar studies with other Drosophila variable exons. Transgenic flies expressing the indirect flight isoform-exon 7a have normal indirect flight muscle structure, and flight and jump ability. However, expression of the embryonic-exon 7d chimeric isoform yields flightless flies that show improvements in both the structural stability of the indirect flight muscle and in locomotor abilities as compared to flies expressing the embryonic isoform. Overall, our results suggest the exon 7 domain participates in the regulation of the attachment of myosin to actin in order to fine-tune the physiological properties of Drosophila myosin isoforms.
Miller
Becky M
BM
Department of Biology and Molecular Biology Institute, San Diego State University, San Diego, CA 92182-4614, USA.
Zhang
Shuxing
S
Suggs
Jennifer A
JA
Swank
Douglas M
DM
Littlefield
Kimberly P
KP
Knowles
Aileen F
AF
Bernstein
Sanford I
SI
D000199:Actins / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001665:Binding Sites; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism*; D005091:Exons / Q000235:genetics; D005426:Flight, Animal; D007700:Kinetics; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009043:Motor Activity; D009132:Muscles / Q000378:metabolism* / Q000648:ultrastructure; D009218:Myosins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D009711:Nucleotides / Q000378:metabolism*; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
D000199:Actins; D009711:Nucleotides; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D000251:Adenosine Triphosphatases; D009218:Myosins