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Inactivation of zebrafish mrf4 leads to myofibril misalignment and motor axon growth disorganization
Author(s) -
Wang YunHsin,
Li ChunKai,
Lee GangHui,
Tsay HueyJen,
Tsai HuaiJen,
Chen YauHung
Publication year - 2008
Publication title -
developmental dynamics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.634
H-Index - 141
eISSN - 1097-0177
pISSN - 1058-8388
DOI - 10.1002/dvdy.21478
Subject(s) - biology , gene knockdown , microbiology and biotechnology , genetics , gene
Mrf4 is a basic helix‐loop‐helix (bHLH) transcription factor associated with myogenesis. Two mrf4 transcripts, mrf4_tv1 and mrf4_tv2 , were identified in zebrafish generated by alternative splicing. To study their biological functions, we separately injected the Mrf4 ‐morpholinos, including MO1 ( mrf4_tv1:mrf4_tv2 knockdown), MO2+MO3 ( mrf4_tv1:mrf4_tv2 knockdown), MO3 ( mrf4_tv1 knockdown), and MO4 ( mrf4_tv2 knockdown), into zebrafish embryos to observe mrf4 gene knockdown phenotypes. No phenotypic abnormalities were observed following injection with 0.5 ng of MO1 but those injected with 4.5, 9, or 13.5 ng displayed curved‐body phenotypes, such as indistinct somite boundaries, and a lack of uniformly sized cell blocks. Similar results were also observed in the (MO2+MO3)‐, MO3‐, and MO4‐injected groups. To further investigate the molecular mechanisms that lead to curved‐body phenotypes, we stained embryos with α‐bungrotoxin and specific monoclonal antibodies F59, Znp1, and Zn5 to detect morphological changes in acetyl‐choline receptor (AChR) clusters, muscle fibers, common path of the primary neurons, and secondary neurons axonal projections, respectively. Our results show that the muscle fibers of mrf4 _ ( tv1:tv2 )‐morphant aligned disorderly and lost their integrity and attachment, while the defects became milder in either mrf4_tv1 ‐morphant or mrf4_tv2 ‐morphant. On the other hand, reduced axonal projections and AChR clusters were found in both mrf4_tv2 ‐morphant and mrf4 _ ( tv1:tv2 )‐morphant but distributed normally in the mrf4_tv1 ‐morphant. We conclude that Mrf4_tv2 is involved in alignment of muscle fibers, and Mrf4_tv1 might have cooperative function with Mrf4_tv2 in muscle fiber alignment, without affecting the muscle‐nerve connection. Developmental Dynamics 237:1043–1050, 2008. © 2008 Wiley‐Liss, Inc.