Lack of WDR36 leads to preimplantation embryonic lethality in mice and delays the formation of small subunit ribosomal RNA in human cells in vitro

Abstract

Mutations in WD repeat domain 36 gene (WDR36) play a causative role in some forms of primary open-angle glaucoma, a leading cause of blindness worldwide. WDR36 is characterized by the presence of multiple WD40 repeats and shows homology to Utp21, an essential protein component of the yeast small subunit (SSU) processome required for maturation of 18S rRNA. To clarify the functional role of WDR36 in the mammalian organism, we generated and investigated mutant mice with a targeted deletion of Wdr36. In parallel experiments, we used RNA interference to deplete WDR36 mRNA in mouse embryos and cultured human trabecular meshwork (HTM-N) cells. Deletion of Wdr36 in the mouse caused preimplantation embryonic lethality, and essentially similar effects were observed when WDR36 mRNA was depleted in mouse embryos by RNA interference. Depletion of WDR36 mRNA in HTM-N cells caused apoptotic cell death and upregulation of mRNA for BAX, TP53 and CDKN1A. By immunocytochemistry, staining for WDR36 was observed in the nucleolus of cells, which co-localized with that of nucleolar proteins such as nucleophosmin and PWP2. In addition, recombinant and epitope-tagged WDR36 localized to the nucleolus of HTM-N cells. By northern blot analysis, a substantial decrease in 21S rRNA, the precursor of 18S rRNA, was observed following knockdown of WDR36. In addition, metabolic-labeling experiments consistently showed a delay of 18S rRNA maturation in WDR36-depleted cells. Our results provide evidence that WDR36 is an essential protein in mammalian cells which is involved in the nucleolar processing of SSU 18S rRNA.

WDR36 acts as a scaffold protein tethering a G-protein-coupled receptor, Gαq and phospholipase Cβ in a signalling complex

“…WDR36 loss of function results in reduced levels of 18S rRNA in zebrafish (Skarie and Link, 2008). Knockdown of WDR36 expression causes a delay in processing 18S rRNA in mammalian cells, and loss of WDR36 in mice leads to preimplantation embryonic lethality (Gallenberger et al, 2011). Chi et al reported that mutant WDR36 directly affects axon growth of retinal ganglion cells in transgenic mice (Chi et al, 2010).…”

Integrative structural analysis of the UTPB complex, an early assembly factor for eukaryotic small ribosomal subunits

“…All six UTPB proteins are universally conserved in eukaryotes and some have been associated with human diseases. For example, mutations of the human UTP21 gene are the causative agents of some forms of primary open angle glaucoma (20) and heterozygous deletion of the UTP6 gene is a candidate modifier of neurofibromatosis type 1 (21). …”

Major review: Molecular genetics of primary open-angle glaucoma

“…However, subsequent studies have failed to replicate these findings. More recent studies suggest that genetic variants of WDR36 may alter POAG risk (Blanco-Marchite et al, 2011; Fan et al, 2009; Fingert et al, 2007; Frezzotti et al, 2011; Gallenberger et al, 2011; Hauser et al, 2006a; Hewitt et al, 2006; Janssen et al, 2013; Kramer et al, 2006; Miyazawa et al, 2007; Mookherjee et al, 2011; Pasutto et al, 2008; Ramdas et al, 2011a; Weisschuh et al, 2007). Knockdown of Wdr36 in zebrafish reportedly reduces levels of 18S rRNA, activates the p53 stress-response pathway, and is associated with ocular abnormalities (Skarie and Link, 2008).…”