Dataset Information

?-Actinin-2 deficiency results in sarcomeric defects in zebrafish that cannot be rescued by ?-actinin-3 revealing functional differences between sarcomeric isoforms.

ABSTRACT: ?-Actinins are actin-binding proteins that can be broadly divided into Ca(2+)-sensitive cytoskeletal and Ca(2+)-insensitive sarcomeric isoforms. To date, little is known about functional differences between the isoforms due to their indistinguishable activities in most in vitro assays. To identify functional differences in vivo between sarcomeric isoforms, we employed computational and molecular approaches to characterize the zebrafish (Danio rerio) genome, which contains orthologoues of each human ?-actinin gene, including duplicated copies of actn3. Each isoform exhibits a distinct and unique pattern of gene expression as assessed by mRNA in situ hybridization, largely sharing similar expression profiles as seen in humans. The spatial conservation of expression of these genes from lower invertebrates to humans suggests that regulation and subsequent functions of these genes are conserved during evolution. Morpholino-based knockdown of the sarcomeric isoform, actn2, leads to skeletal muscle, cardiac, and ocular defects evident over the first week of development. Remarkably, despite the high degree of sequence conservation between actn2 and actn3, the phenotypes of ?-actinin-2 deficient zebrafish can be rescued by overexpression of ?-actinin-2 but not by ?-actinin-3 mRNAs from zebrafish or human. These data provide functional evidence that the primary sequences of ?-actinin-2 and ?-actinin-3 evolved differences to optimize their functions.

SUBMITTER: Gupta V

PROVIDER: S-EPMC3336783 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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α-Actinin-2 deficiency results in sarcomeric defects in zebrafish that cannot be rescued by α-actinin-3 revealing functional differences between sarcomeric isoforms.

Gupta Vandana V Discenza Marie M Guyon Jeffrey R JR Kunkel Louis M LM Beggs Alan H AH

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20120117 5

α-Actinins are actin-binding proteins that can be broadly divided into Ca(2+)-sensitive cytoskeletal and Ca(2+)-insensitive sarcomeric isoforms. To date, little is known about functional differences between the isoforms due to their indistinguishable activities in most in vitro assays. To identify functional differences in vivo between sarcomeric isoforms, we employed computational and molecular approaches to characterize the zebrafish (Danio rerio) genome, which contains orthologoues of each hu ...[more]

PMID: 22253474

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Project description:Primary steps in vertebrate vision occur in rod and cone cells of the retina and require precise molecular switches in excitation, recovery, and adaptation. In particular, recovery of the photoresponse and light adaptation processes are under control of neuronal Ca2+ sensor (NCS) proteins. Among them, the Ca2+ sensor recoverin undergoes a pronounced Ca2+-dependent conformational change, a prototypical so-called Ca2+-myristoyl switch, which allows selective targeting of G protein-coupled receptor kinase. Zebrafish (Danio rerio) has gained attention as a model organism in vision research. It expresses four different recoverin isoforms (zRec1a, zRec1b, zRec2a, and zRec2b) that are orthologs to the one known mammalian variant. The expression pattern of the four isoforms cover both rod and cone cells, but the differential distribution in cones points to versatile functions of recoverin in these cell types. Initial functional studies on zebrafish larvae indicate different Ca2+-sensitive working modes for zebrafish recoverins, but experimental evidence is lacking so far. The aims of the present study are (1) to measure specific Ca2+-sensing properties of the different recoverin isoforms, (2) to ask whether switch mechanisms triggered by Ca2+ resemble that one observed with mammalian recoverin, and (3) to investigate a possible impact of an attached myristoyl moiety. For addressing these questions, we employ fluorescence spectroscopy, surface plasmon resonance (SPR), dynamic light scattering, and equilibrium centrifugation. Exposure of hydrophobic amino acids, due to the myristoyl switch, differed among isoforms and depended also on the myristoylation state of the particular recoverin. Ca2+-induced rearrangement of the protein-water shell was for all variants less pronounced than for the bovine ortholog indicating either a modified Ca2+-myristoyl switch or no switch. Our results have implications for a step-by-step response of recoverin isoforms to changing intracellular Ca2+ during illumination.

Dataset Information

?-Actinin-2 deficiency results in sarcomeric defects in zebrafish that cannot be rescued by ?-actinin-3 revealing functional differences between sarcomeric isoforms.

Publications

α-Actinin-2 deficiency results in sarcomeric defects in zebrafish that cannot be rescued by α-actinin-3 revealing functional differences between sarcomeric isoforms.

Similar Datasets

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