Zebrafish Tric-b is required for skeletal development and bone cells differentiation

Abstract

Introduction: Trimeric intracellular potassium channels TRIC-A and -B are endoplasmic reticulum (ER) integral membrane proteins, involved in the regulation of calcium release mediated by ryanodine (RyRs) and inositol 1,4,5-trisphosphate (IP₃Rs) receptors, respectively. While TRIC-A is mainly expressed in excitable cells, TRIC-B is ubiquitously distributed at moderate level. TRIC-B deficiency causes a dysregulation of calcium flux from the ER, which impacts on multiple collagen specific chaperones and modifying enzymatic activity, leading to a rare form of osteogenesis imperfecta (OI Type XIV). The relevance of TRIC-B on cell homeostasis and the molecular mechanism behind the disease are still unknown.

Results: In this study, we exploited zebrafish to elucidate the role of TRIC-B in skeletal tissue. We demonstrated, for the first time, that tmem38a and tmem38b genes encoding Tric-a and -b, respectively are expressed at early developmental stages in zebrafish, but only the latter has a maternal expression. Two zebrafish mutants for tmem38b were generated by CRISPR/Cas9, one carrying an out of frame mutation introducing a premature stop codon (tmem38b^-/- ) and one with an in frame deletion that removes the highly conserved KEV domain (tmem38b^{Δ120-7/Δ120-7} ). In both models collagen type I is under-modified and partially intracellularly retained in the endoplasmic reticulum, as described in individuals affected by OI type XIV. Tmem38b^-/- showed a mild skeletal phenotype at the late larval and juvenile stages of development whereas tmem38b^{Δ120-7/Δ120-7} bone outcome was limited to a reduced vertebral length at 21 dpf. A caudal fin regeneration study pointed towards impaired activity of osteoblasts and osteoclasts associated with mineralization impairment.

Discussion: Our data support the requirement of Tric-b during early development and for bone cell differentiation.

Keywords: TRIC-B; bone; collagen; endoplasmic reticulum; fin regeneration; osteogenesis imperfecta; zebrafish.