Zebrafish Collagen XIV Is Transiently Expressed in Epithelia and Is Required for Proper Function of Certain Basement Membranes

Hannah L Bader,Elise Lambert,Alexandre Guiraud,Marilyne Malbouyres,Wolfgang Driever,Manuel Koch,Florence Ruggiero

doi:10.1074/jbc.m112.430637

Abstract

We found that zebrafish has two differentially expressed col14a1 paralogs. col14a1a expression peaked between 18-somite stage and 24 hours postfertilization (hpf), whereas col14a1b was first expressed at 32 hpf. To uncover functions of collagen XIV (COLXIV) during early embryogenesis, we focused our study on col14a1a. We characterized the α1 (XIV-A) chain as a collagenase-sensitive 200-kDa protein that formed dimer that could be reduced at high pH. As observed for the transcript, COLXIV-A protein expression peaked between 24 and 48 hpf. Using antisense probes and polyclonal antibodies, we show that col14a1a and its protein product COLXIV-A are transiently expressed in several epithelia, including epithelia undergoing shape changes, such as the fin folds. In contrast, anti-COLXII antibodies stained only connective tissues. COLXIV-A was also detected in the basement membrane (BM), where it co-localized with COLXII. At later developmental stages, COLXIV-A was not expressed in epithelia anymore but persisted in the BM. Morpholino knockdown of COLXIV-A provoked a skin detachment phenotype. Electron microscopy analysis revealed that morpholino-injected embryos lacked a lamina densa and lamina lucida at 24 hpf, and BM defects, such as gaps in the adepidermal granules, were still detected at 48 hpf. These BM defects were accompanied by a rupture of the dermis and detachment of the epidermis. Taken together, these data suggest an unexpected role of COLXIV-A in undifferentiated epithelia and in the formation of embryonic basement membranes.

Highlights

Collagen XIV (COLXIV) is a component of connective tissues and regulates type I collagen fibril assembly in mice
COLXIV-A was detected in the basement membrane (BM), where it co-localized with COLXII
Using zebrafish anti-COLXII [14] and newly prepared anti-COLXIV-A antibodies, we found marked differences in the expression pattern of these two collagens: COLXII was expressed in connective tissues [14], whereas COLXIV-A was expressed in epithelia

Summary

Background

Collagen XIV (COLXIV) is a component of connective tissues and regulates type I collagen fibril assembly in mice. Using zebrafish anti-COLXII [14] and newly prepared anti-COLXIV-A antibodies, we found marked differences in the expression pattern of these two collagens: COLXII was expressed in connective tissues [14], whereas COLXIV-A was expressed in epithelia Their expression overlapped in the basement membrane and in particular in the epi-. Morpholino knockdown of COLXIV-A demonstrated a crucial role of this protein in the proper formation of the epidermal-dermal basement membrane and in the cohesion of the epidermis and the subjacent dermis. We demonstrate that 1) COLXIV-A is embryonically regulated, 2) collagens XII and XIV may have distinct spatiotemporal expression patterns during development, and 3) at early developmental stages, COLXIV-A has a role in undifferentiated epithelia and in the formation of the basement membrane

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION