Collagen XV, a novel factor in zebrafish notochord differentiation and muscle development

Abstract

Muscle cells are surrounded by extracellular matrix, the components of which play an important role in signalling mechanisms involved in their development. In mice, loss of collagen XV, a component of basement membranes expressed primarily in skeletal muscles, results in a mild skeletal myopathy. We have determined the complete zebrafish collagen XV primary sequence and analysed its expression and function in embryogenesis. During the segmentation period, expression of the Col15a1 gene is mainly found in the notochord and its protein product is deposited exclusively in the peri-notochordal basement membrane. Morpholino mediated knock-down of Col15a1 causes defects in notochord differentiation and in fast and slow muscle formation as shown by persistence of axial mesodermal marker gene expression, disorganization of the peri-notochodal basement membrane and myofibrils, and a U-shape myotome. In addition, the number of medial fast-twitch muscle fibers was substantially increased, suggesting that the signalling by notochord derived Hh proteins is enhanced by loss of collagen XV. Consistent with this, there is a concomitant expansion of patched-1 expression in the myotome of morphant embryos. Together, these results indicate that collagen XV is required for notochord differentiation and muscle development in the zebrafish embryo and that it interplays with Shh signalling.

Characterization of drCol 15a1b: A Novel Component of the Stem Cell Niche in the Zebrafish Retina

“…Bioinformatic analyses indicated that this EST was most similar to a region of the collagen alpha-1 XV chain precursor (col 15a1) located in linkage group 2 (ENSEMBL database ENSDARG000000 61848, release 50). The sequence of the EST is, however, substantially different from that recently published for zebrafish col 15a1 [26], indicating that the EST-match represents for a novel duplicate of col 15a1 gene in the zebrafish, which we have named ''drCol 15a1b'' (to distinguish it from that previously described by [26], which we will now refer to as ''drCol 15a1a''). Comparison with the EST sequence did not fully characterize the open reading frame of drCol 15a1b, since it did not include the translation startpoint.…”

“…As mentioned previously, the multiplexin family of collagens comprises the collagen XV (col 15a1) and XVIII (col 18a1), that in the case of zebrafish presents two duplicate sequences for each of the two genes: drCol 15a1a [26], drCol 15a1b (the one studied here), drCol 18a1 [27], and a novel sequence with high homology with col 18a1 (database access number ENSDARG00000041746), which has been discovered when searching the TreeFam database. Expression studies have revealed that drCol 15a1b shows a different expression pattern from those reported for drCol 15a1a and for drCol 18a1 in the zebrafish.…”

“…We have characterized a new gene from the zebrafish that codes for a collagen alpha-1 XV chain precursor that is different from the previously reported one [26]. Therefore, we have used the term ''drCol 15a1a'' for the previously identified gene [26] and ''drCol 15a1b'' for the one reported here by us.…”

“…Therefore, we have used the term ''drCol 15a1a'' for the previously identified gene [26] and ''drCol 15a1b'' for the one reported here by us. Bioinformatic analyses indicate that part of the sequence from drCol 15a1a aligns with genomic clones from linkage group 24 and the complete open reading frame of drCol 15a1b is located in linkage group 2.…”

“…The complete open reading frame codes a protein of 1,236 residues, which displays the common features observed for this protein family, that is, a signal peptide, a thrombospondin domain, five collagen helical domains and a C-terminal endostatin peptide (Supporting Information Figures S1 and S2). drCol 15a1b protein has 48% identity at the amino acid level with its duplicate, drCol 15a1a [26], 43% with its human ortholog, and 34% and 40% of identity with its paralogs col 18a1 in the zebrafish and human, respectively.…”