Deficiency of a Transmembrane Prolyl 4-Hydroxylase in the Zebrafish Leads to Basement Membrane Defects and Compromised Kidney Function

Prolyl 4-hydroxylases (P4Hs) catalyze the hydroxylation of collagens and hypoxia-inducible factor (HIF)-alpha subunits. We studied the zebrafish homologue of the recently characterized human transmembrane P4H (P4H-TM) that can hydroxylate HIF-alpha, but not collagens, in vitro and influence HIF-alpha levels in cellulo. The zebrafish P4H-TM mRNA had its highest expression in the eye and brain and lower levels in other tissues, including the kidney. Morpholino knockdown of P4H-TM in embryos resulted in a reduction in the size of the eye and head and morphological alterations in the head from 2 days postfertilization onward. In addition, pericardial edema, regarded as a sign of kidney dysfunction, developed from 3 days postfertilization onward. The phenotype was dependent on the P4H-TM catalytic activity because similar results were obtained with morpholinos targeting either translation initiation or catalytic residues of the enzyme. Structural and functional analyses of the morphant pronephric kidneys revealed fragmented glomerular basement membranes (BMs), disorganized podocyte foot processes, and severely compromised pronephric kidney function leading to proteinuria. The opacity of the eye lens was increased due to the presence of extra nuclei and deposits, and the structure of the lens capsule BM was altered.