Tetraspanin Cd9b and Cxcl12a/Cxcr4b have a synergistic effect on the control of collective cell migration

Abstract

Collective cell migration is essential for embryonic development and homeostatic processes. During zebrafish development, the posterior lateral line primordium (pLLP) navigates along the embryo flank by collective cell migration. The chemokine receptors, Cxcr4b and Cxcr7b, as well as their cognate ligand, Cxcl12a, are essential for this process. We corroborate that knockdown of the zebrafish cd9 tetraspanin orthologue, cd9b, results in mild pLL abnormalities. Through generation of CRISPR and TALEN mutants, we show that cd9a and cd9b function partially redundantly in pLLP migration, which is delayed in the cd9b single and cd9a; cd9b double mutants. This delay led to a transient reduction in neuromast numbers. Loss of both Cd9a and Cd9b sensitized embryos to reduced Cxcr4b and Cxcl12a levels. Together these results provide evidence that Cd9 modulates collective cell migration of the pLLP during zebrafish development. One interpretation of these observations is that Cd9 contributes to more effective chemokine signalling.

Tetraspanin Cd9b plays a role in fertility in zebrafish

“…While both mutations occur in similar codons (15 and 16), the consequent frameshift causes slightly more aberrant amino acids in the cd9b is16 KOs before a stop codon is created (46 in contrast to 22 in cd9b pg15 KO). While ISH and qPCR results showed a downregulation of mRNA suggesting nonsense mediated decay was occurring ( S2 , S3 Figs) [ 15 ], there could be differences in the residual function of the mutated protein. It is known, for example, that synthetic cell permeable peptides corresponding to the termini of CD9 show cellular activity [ 18 ].…”

“…This increases the likelihood that there is some compensation happening in the cd9b KOs. The most likely candidate is cd9a as they share high protein identity and similar mRNA expression patterns to cd9b [ 15 ].…”

“…To determine if deletion of both paralogs would result in a complete loss of fertility as seen in the CD9 KO mouse, cd9a was knocked out in the cd9b pg15 background. ISH results show downregulation of cd9b and cd9a mRNA suggesting nonsense mediated decay was occurring [ 15 ]. Interestingly, the fertilisation rate of the double KO line was very similar to the cd9b pg15 KO line ( Fig 1D ), suggesting that only Cd9b is involved in egg production and fertilisation.…”

“…Tetraspanins are widely expressed in teleosts [ 9 ] but there are no reports of roles in fish fertility. Several tetraspanins have roles during zebrafish development, including pigment cell interactions [ 10 ], hatching [ 11 ], vascularisation [ 12 , 13 ] migrasome formation [ 14 ] and primordium migration [ 15 ]. However, the role of Cd9 in fertilisation has not yet been investigated.…”

“…However, the role of Cd9 in fertilisation has not yet been investigated. There are two paralogues of cd9 in zebrafish (cd9a and cd9b), which have 63% amino acid identity and similar mRNA expression patterns [ 15 ].…”