Endocytic recycling protein EHD1 regulates primary cilia morphogenesis and SHH signaling during neural tube development

Abstract

Members of the four-member C-terminal EPS15-Homology Domain-containing (EHD) protein family play crucial roles in endocytic recycling of cell surface receptors from endosomes to the plasma membrane. In this study, we show that Ehd1 gene knockout in mice on a predominantly B6 background is embryonic lethal. Ehd1-null embryos die at mid-gestation with a failure to complete key developmental processes including neural tube closure, axial turning and patterning of the neural tube. We found that Ehd1-null embryos display short and stubby cilia on the developing neuroepithelium at embryonic day 9.5 (E9.5). Loss of EHD1 also deregulates the ciliary SHH signaling with Ehd1-null embryos displaying features indicative of increased SHH signaling, including a significant downregulation in the formation of the GLI3 repressor and increase in the ventral neuronal markers specified by SHH. Using Ehd1-null MEFS we found that EHD1 protein co-localizes with the SHH receptor Smoothened in the primary cilia upon ligand stimulation. Under the same conditions, EHD1 was shown to co-traffic with Smoothened into the developing primary cilia and we identify EHD1 as a direct binding partner of Smoothened. Overall, our studies identify the endocytic recycling regulator EHD1 as a novel regulator of the primary cilium-associated trafficking of Smoothened and Hedgehog signaling.

CSF-1 receptor signalling is governed by pre-requisite EHD1 mediated receptor display on the macrophage cell surface

“…Ehd1 fl/fl mice in a predominantly C57BL/6 background [37,38] were crossed with tamoxifen-inducible CreERT2 expressing mice from Jackson Laboratories ( Gt(ROSA)26Sor tm1(cre/ERT2)Tyj ; strain 008463) to generate Ehd1 fl/fl ; Cre ERT2 mice. Genotypes were confirmed by subjecting tail clip DNA to PCR analysis using the KAPA mouse genotyping kit (KAPA Biosystems) and primers pairs described previously [36,38].…”

“…We have found that EHD1 deletion on a mixed C57BL/6 and 129Sv background is partially penetrant embryonic lethal and associated with defective spermatogenesis and lens developmental defects, while it is fully embryonic lethal on a predominantly C57BL/6 background [35–37]. Here, we use primary bone marrow-derived macrophages (BMDMs) with inducible EHD1-Knockout (EHD1-KO) capacity, to study the biological and physiological function of EHD1 in macrophages.…”

Retromer associates with the cytoplasmic amino-terminus of polycystin-2

“…Exocyst, for example, was initially characterised as being required for the vesicular transport of cargo proteins from the Golgi complex to the plasma membrane in yeast (Novick et al, 1980;TerBush et al, 1996). Members of the Eps15 homology domain (EHD) protein family, which have roles in early stages of ciliogenesis, have long been known to function in membrane remodelling processes within the endosomal network (Naslavsky and Caplan, 2011;Lu et al, 2015;Bhattacharyya et al, 2016). Additionally, a recent study has demonstrated localisation of the endosomal protein serologically defined colon cancer antigen-3 (SDCCAG3) to the basal body of primary cilia, and implicated its activity in ciliary targeting of PC2 (Yu et al, 2016).…”

The regulation of cilium assembly and disassembly in development and disease

“…3). Although the cues are not fully understood, a sequence of events, triggered by Rab11 and Rabin8 (also known as RAB3IP), promotes the recruitment of the small GTPase Rab8a to CV to facilitate their extension (Bhattacharyya et al, 2016;Lu et al, 2015b;Wu et al, 2018). Rab8a facilitates the docking of PCVs to DAs by interacting with a group of distal centriolar proteins, including CEP164, Chibby (also known as CBY1), AHI1 and Talpid3 (also known as KIAA0586) (Burke et al, 2014;Hsiao et al, 2009;Kobayashi et al, 2014;Schmidt et al, 2012).…”