Human coronavirus 3CL proteases cleave septins and disrupt Hedgehog signaling, causing ciliary dysfunction.

Coronaviruses target ciliate cells causing the loss of cilia, acute rhinorrheas, and other ciliopathies. The loss of ciliary function may help the virus infect, replicate, and spread. However, the molecular mechanisms by which coronaviruses cause ciliary defects are still unclear. Herein we demonstrate how coronavirus infection and severe acute respiratory syndrome coronavirus2 3CL protease induce cilia dysfunction by targeting a host protein septin that is required for the structure and function of cilia. Further, we demonstrate that coronaviruses and 3CL protease lead to the cleavage of several septin proteins (SEPT2, -6, and -9), producing cleaved obstructive fragments. Furthermore, ectopic expression of cleaved SEPT2 fragments shows defective ciliogenesis, disoriented septin filaments, and ablated Sonic Hedgehog (SHH) signaling in a protease activity-dependent manner. We present that the 3CLpro inhibitors are potent and prevent abnormal ciliary structures and SHH signaling. These results provide useful insights into the general mechanisms underlying ciliary defects caused by coronaviruses, which, in turn, facilitate virus spread and prove that preclinical and clinical 3CL protease inhibitors may prove useful as therapeutics for treating ciliary defects of coronaviruses.

GJA1 depletion causes ciliary defects by affecting Rab11 trafficking to the ciliary base.

The gap junction complex functions as a transport channel across the membrane. Among gap junction subunits, gap junction protein α1 (GJA1) is the most commonly expressed subunit. A recent study showed that GJA1 is necessary for the maintenance of motile cilia; however, the molecular mechanism and function of GJA1 in ciliogenesis remain unknown. Here, we examined the functions of GJA1 during ciliogenesis in human retinal pigment epithelium-1 and Xenopus laevis embryonic multiciliated-cells. GJA1 localizes to the motile ciliary axonemes or pericentriolar regions beneath the primary cilium. GJA1 depletion caused malformation of both the primary cilium and motile cilia. Further study revealed that GJA1 depletion affected several ciliary proteins such as BBS4, CP110, and Rab11 in the pericentriolar region and basal body. Interestingly, CP110 removal from the mother centriole was significantly reduced by GJA1 depletion. Importantly, Rab11, a key regulator during ciliogenesis, was immunoprecipitated with GJA1 and GJA1 knockdown caused the mislocalization of Rab11. These findings suggest that GJA1 regulates ciliogenesis by interacting with the Rab11-Rab8 ciliary trafficking pathway.

Metastasis enhancer PGRMC1 boosts store-operated Ca2+ entry by uncoiling Ca2+ sensor STIM1 for focal adhesion turnover and actomyosin formation.

Progesterone receptor membrane component 1 (PGRMC1), the overexpression of which reduces survivability of cancer patients, is essential for cell migration and metastasis. However, the intracellular signaling pathways involved are largely unknown. Here, we report that PGRMC1 promotes store-operated Ca2+ entry (SOCE) as a functional interactor of stromal interaction molecule 1 (STIM1). PGRMC1 was repeatedly detected as an interactor of STIM1-Orai1 complex via complementation-dependent in situ labeling. Genetic depletion of PGRMC1 decreased SOCE and impaired activation of the nuclear factor of the activated T cell (NFAT) pathway. Mechanistically, PGRMC1 directly bound to the coiled-coil domain of STIM1, promoting STIM1 conformational switch. In breast cancer cells, PGRMC1 depletion reduced epidermal growth factor (EGF)-induced SOCE and disrupted focal adhesion turnover and actomyosin formation. These findings identify PGRMC1 as an essential regulator of Ca2+ signaling in breast cancer cells, providing a target for treating cancer metastasis and an insight for dissecting various PGRMC1/SOCE-induced biological processes.

Distinct gating mechanism of SOC channel involving STIM-Orai coupling and an intramolecular interaction of Orai in Caenorhabditis elegans.

Store-operated calcium entry (SOCE), an important mechanism of Ca2+ signaling in a wide range of cell types, is mediated by stromal interaction molecule (STIM), which senses the depletion of endoplasmic reticulum Ca2+ stores and binds and activates Orai channels in the plasma membrane. This inside-out mechanism of Ca2+ signaling raises an interesting question about the evolution of SOCE: How did these two proteins existing in different cellular compartments evolve to interact with each other? We investigated the gating mechanism of Caenorhabditis elegans Orai channels. Our analysis revealed a mechanism of Orai gating by STIM binding to the intracellular 2-3 loop of Orai in C. elegans that is radically different from Orai gating by STIM binding to the N and C termini of Orai in mammals. In addition, we found that the conserved hydrophobic amino acids in the 2-3 loop of Orai1 are important for the oligomerization and gating of channels and are regulated via an intramolecular interaction mechanism mediated by the N and C termini of Orai1. This study identifies a previously unknown SOCE mechanism in C. elegans and suggests that, while the STIM-Orai interaction is conserved between invertebrates and mammals, the gating mechanism for Orai channels differs considerably.