The Golgi matrix protein giantin is required for normal cilia function in zebrafish

Abstract

The Golgi is essential for glycosylation of newly synthesised proteins including almost all cell-surface and extracellular matrix proteoglycans. Giantin, encoded by the golgb1 gene, is a member of the golgin family of proteins that reside within the Golgi stack, but its function remains elusive. Loss of function of giantin in rats causes osteochondrodysplasia; knockout mice show milder defects, notably a cleft palate. In vitro, giantin has been implicated in Golgi organisation, biosynthetic trafficking, and ciliogenesis. Here we show that loss of function of giantin in zebrafish, using either morpholino or knockout techniques, causes defects in cilia function. Giantin morphants have fewer cilia in the neural tube and those remaining are longer. Mutants have the same number of cilia in the neural tube but these cilia are also elongated. Scanning electron microscopy shows that loss of giantin results in an accumulation of material at the ciliary tip, consistent with a loss of function of retrograde intraflagellar transport. Mutants show milder defects than morphants consistent with adaptation to loss of giantin.

Regulator of calcineurin-2 is a centriolar protein with a role in cilia length control

“…Dynein-2 is the major motor driving retrograde IFT along the axoneme and is, among other functions, required for the transduction of the Shh signal (He et al, 2016). Consistent with our findings using RNAi in cultured cells, morpholino knockdown of giantin in zebrafish resulted in fewer but longer cilia in the neural tube (Bergen et al, 2017). In contrast, recent characterisation of giantin KO zebrafish shows that, while breeding and developing grossly similar to wild-type fish, giantin knockout (KO) fish show a significant decrease in body length (more notable in young adults) and have defects in extracellular matrix, cartilage and bone formation, but only minor cilia defects (Bergen et al, 2017).…”

“…Giantin-depleted cells produce fewer cilia and those that remain are longer suggesting a defect in length control. In contrast, analysis of giantin knockout RPE-1 cells (Stevenson et al, 2017) have no gross defects in ciliogenesis at the level of light microscopy ( Fig. 1A, quantified in Bi) or any significant change in cilia length (Fig.…”

“…1C, enlarged in Cii). Furthermore, Golgi structure appears normal (see (Stevenson et al, 2017)). This phenotypic discrepancy between knockdown and KO cells is consistent with our observations in zebrafish (Bergen et al, 2017) and led us to hypothesise that altered expression of another gene or genes compensates for loss of giantin.…”

“…These data are described in Stevenson et al (2017) and the raw data are available via the ArrayExpress database (www.ebi.ac.uk/arrayexpress) under accession number E-MTAB-5618. The most highly upregulated gene, and therefore the most likely to underpin any compensation for loss of giantin, is RCAN2.…”

Meeting report – Cellular dynamics: membrane–cytoskeleton interface

“…For example, David Stephens (University of Bristol, Bristol, UK) showed that loss of giantin (GOLGB1), but not of other Golgi components, causes defects in cilia formation and a swelling at the tip of cilia due to disrupted retrograde transport. Giantin loss depletes dynein 2 and its regulators from the cilium (Bergen et al, 2017). Last but not least, David Glover (University of Cambridge, Cambridge, UK) presented another link between the Golgi and cilium as he described how the Golgi protein GORAB interacts with the core centriole protein SASS6 and affects the nine-fold symmetry of the centriole in cilia.…”