Trafficking of integral membrane proteins to cilia is poorly understood. Badgandi et al. show that tubby family proteins TULP3 and TUB act as general adapters for ciliary trafficking of structurally diverse integral membrane cargo like GCPRs and the polycystin 1/2 complex.

Pal et al. describe a two-step process determining removal of the cilia-localized GPCR, Gpr161, upon sonic hedgehog signaling. First, β-arrestins are recruited by the signaling-competent receptor in a smoothened-dependent manner. Second, clathrin-mediated endocytosis outside of the ciliary compartment coordinates removal.

SUMMARY The eight-subunit T cell receptor (TCR)-CD3 complex is the primary determinant for T cell fate decisions. Yet how it relays ligand-specific information across the cell membrane for conversion to chemical signals remains unresolved. We hypothesized that TCR engagement triggers a change in the spatial relationship between the associated CD3ζζ subunits at the junction where they emerge from the membrane into the cytoplasm. Using three in situ proximity assays based on ID-PRIME, FRET, and EPOR activity we determined that the cytosolic juxtamembrane regions of the CD3ζζ subunits are spread apart upon assembly into the TCR-CD3 complex. TCR engagement then triggered their apposition. This mechanical switch resides upstream of the CD3ζζ intracellular motifs that initiate chemical signaling as well as the polybasic stretches that regulate signal potentiation. These findings provide a framework from which to examine triggering events for activating immune receptors and other complex molecular machines.

Highlights d Nephron-specific Tulp3 knockouts develop cystic kidneys without disrupting cilia d Cystogenesis is intermediate between that caused by loss of polycystin-1 or cilia d Polycystin-2 and Arl13b are reduced in cilia of knockout collecting duct cells d Arl13b is the likely polycystin-independent ciliary factor repressing cystogenesis

The pre-T-cell receptor (TCR)-, αβTCR-, and γδTCR-CD3 complexes are members of a family of modular biosensors that are responsible for driving T-cell development, activation, and effector functions. They inform essential checkpoint decisions by relaying key information from their ligand-binding modules (TCRs) to their signaling modules (CD3γε + CD3δε and CD3ζζ) and on to the intracellular signaling apparatus. Their actions shape the T-cell repertoire, as well as T-cell-mediated immunity; yet, the mechanisms that underlie their activity remain an enigma. As with any molecular machine, understanding how they function depends upon understanding how their parts fit and work together. In the 30 years since the initial biochemical and genetic characterizations of the αβTCR, the structure and function of the individual components of these family members have been extensively characterized. Cumulatively, this information has allowed us to piece together a portrait of the αβTCR-CD3 complex and outline the form of the remaining family members. Here we review the known structural and functional characteristics of the components of these TCR-CD3 complex family members. We then discuss how these data have informed our understanding of the architecture of the αβTCR-CD3 complex as well as their implications for the other family members. The intent is to provide a framework for considering: (i) how these thematically similar complexes diverge to execute their specific functions and (ii) how our knowledge of the form and function of these distinct family members can cross-inform our understanding of the other family members.

The primary cilium has been found to be associated with a number of cellular signaling pathways, such as vertebrate hedgehog signaling, and implicated in the pathogenesis of diseases affecting multiple organs, including the neural tube, kidney, and brain. The primary cilium is the site where a subset of the cell's membrane proteins is enriched. However, pathways that target and concentrate membrane proteins in cilia are not well understood. Processes determining the level of proteins in the ciliary membrane include entry into the compartment, removal, and retention by diffusion barriers such as the transition zone. Proteins that are concentrated in the ciliary membrane are also localized to other cellular sites. Thus it is critical to determine the particular role for ciliary compartmentalization in sensory reception and signaling pathways. Here we provide a brief overview of our current understanding of compartmentalization of proteins in the ciliary membrane and the dynamics of trafficking into and out of the cilium. We also discuss major unanswered questions regarding the role that defects in ciliary compartmentalization might play in disease pathogenesis. Understanding the trafficking mechanisms that underlie the role of ciliary compartmentalization in signaling might provide unique approaches for intervention in progressive ciliopathies.

Blood feeding by vector mosquitoes provides the entry point for disease pathogens and presents an acute metabolic challenge that must be overcome to complete the gonotrophic cycle. Based on recent data showing that coatomer protein I (COPI) vesicle transport is involved in cellular processes beyond Golgi–endoplasmic reticulum retrograde protein trafficking, we disrupted COPI functions in the Yellow Fever mosquito Aedes aegypti to interfere with blood meal digestion. Surprisingly, we found that decreased expression of the γCOPI coatomer protein led to 89% mortality in blood-fed mosquitoes by 72 h postfeeding compared with 0% mortality in control dsRNA-injected blood-fed mosquitoes and 3% mortality in γCOPI dsRNA-injected sugar-fed mosquitoes. Similar results were obtained using dsRNA directed against five other COPI coatomer subunits (α, β, β′, δ, and ζ). We also examined midgut tissues by EM, quantitated heme in fecal samples, and characterized feeding-induced protein expression in midgut, fat body, and ovary tissues of COPI-deficient mosquitoes. We found that COPI defects disrupt epithelial cell membrane integrity, stimulate premature blood meal excretion, and block induced expression of several midgut protease genes. To study the role of COPI transport in ovarian development, we injected γCOPI dsRNA after blood feeding and found that, although blood digestion was normal, follicles in these mosquitoes were significantly smaller by 48 h postinjection and lacked eggshell proteins. Together, these data show that COPI functions are critical to mosquito blood digestion and egg maturation, a finding that could also apply to other blood-feeding arthropod vectors.

The mechanisms underlying subcellular targeting of cAMP-generating adenylyl cyclases and processes regulated by their compartmentalization are poorly understood. Here, we identify Ankmy2 as a repressor of the Hedgehog pathway via adenylyl cyclase targeting. Ankmy2 binds to multiple adenylyl cyclases, determining their maturation and trafficking to primary cilia. Mice lacking Ankmy2 are mid-embryonic lethal. Knockout embryos have increased Hedgehog signaling and completely open neural tubes showing co-expansion of all ventral neuroprogenitor markers, comparable to the loss of the Hedgehog receptor Patched1. Ventralization in Ankmy2 knockout is completely independent of the Hedgehog pathway transducer Smoothened. Instead, ventralization results from the reduced formation of Gli2 and Gli3 repressors and early depletion of adenylyl cyclase III in neuroepithelial cilia, implicating deficient pathway repression. Ventralization in Ankmy2 knockout requires both cilia and Gli2 activation. These findings indicate that cilia-dependent adenylyl cyclase signaling represses the Hedgehog pathway and promotes morphogenetic patterning.