The cilium, a pivotal organelle crucial for cell signaling and proper cell function, relies on meticulous macromolecular transport from the cytoplasm for its formation and maintenance. While the intraflagellar transport (IFT) pathway has traditionally been the focus of extensive study concerning ciliogenesis and ciliary maintenance, recent research highlights a complementary and alternative mechanism-vesicle-assisted transport (VAT) in cytoplasm to cilium trafficking. Despite its potential significance, the VAT pathway remains largely uncharacterized. This review explores recent studies providing evidence for the dynamics of vesicle-related diffusion and transport within the live primary cilium, employing high-speed super-resolution light microscopy. Additionally, we analyze the spatial distribution of vesicles in the cilium, mainly relying on electron microscopy data. By scrutinizing the VAT pathways that facilitate cargo transport into the cilium, with a specific emphasis on recent advancements and imaging data, our objective is to synthesize a comprehensive model of ciliary transport through the integration of IFT-VAT mechanisms.

Lysosomes and lysosome related organelles (LROs) are dynamic organelles at the intersection of various pathways involved in maintaining cellular hemostasis and regulating cellular functions. Vesicle trafficking of lysosomes and LROs are critical to maintain their functions. The lysosomal trafficking regulator (LYST) is an elusive protein important for the regulation of membrane dynamics and intracellular trafficking of lysosomes and LROs. Mutations to the LYST gene result in Chédiak-Higashi syndrome, an autosomal recessive immunodeficiency characterized by defective granule exocytosis, cytotoxicity, etc. Despite eight decades passing since its initial discovery, a comprehensive understanding of LYST\'s function in cellular biology remains unresolved. Accumulating evidence suggests that dysregulation of LYST function also manifests in other disease states. Here, we review the available literature to consolidate available scientific endeavors in relation to LYST and discuss its relevance for immunomodulatory therapies, regenerative medicine and cancer applications.

In B cells, antigen processing and peptide-antigen (pAg) presentation is essential to ignite high-affinity antibody responses with the help of cognate T cells. B cells efficiently internalize and direct specific antigens for processing and loading onto MHCII. This critical step, which enables pAg presentation, occurs in MHCII compartments (MIICs) which possess the enzymatic machinery for pAg loading on MHCII. The intracellular transport systems that guide antigen and maintain this unique compartment remain enigmatic. Here, we probed the possible functional role of two known endosomal proteins, the Rab family small GTPases Rab7 and Rab9, that are both reported to colocalize with internalized antigen. As compared to Rab9, we found Rab7 to exhibit a higher overlap with antigen and MIIC components. Rab7 also showed a higher association with antigen degradation. The inhibition of Rab7 drastically decreased pAg presentation. Additionally, we detected the strong colocalization of perinuclearly clustered and presumably MIIC-associated antigen with autophagy protein LC3. When we pharmacologically inhibited autophagy, pAg presentation was inhibited. Together, our data promote Rab7 as an important regulator of antigen processing and, considering the previously reported functions of Rab7 in autophagy, this also raises the possibility of the involvement of autophagy-related machinery in this process.

Membranes are essential for life. They act as semi-permeable boundaries that define cells and organelles. In addition, their surfaces actively participate in biochemical reaction networks, where they confine proteins, align reaction partners, and directly control enzymatic activities. Membrane-localized reactions shape cellular membranes, define the identity of organelles, compartmentalize biochemical processes, and can even be the source of signaling gradients that originate at the plasma membrane and reach into the cytoplasm and nucleus. The membrane surface is, therefore, an essential platform upon which myriad cellular processes are scaffolded. In this review, we summarize our current understanding of the biophysics and biochemistry of membrane-localized reactions with particular focus on insights derived from reconstituted and cellular systems. We discuss how the interplay of cellular factors results in their self-organization, condensation, assembly, and activity, and the emergent properties derived from them.

The plant endomembrane system is organized and regulated by large gene families that encode proteins responsible for the spatiotemporal delivery and retrieval of cargo throughout the cell and to and from the plasma membrane. Many of these regulatory molecules form functional complexes like the SNAREs, exocyst, and retromer, which are required for the delivery, recycling, and degradation pathways of cellular components. The functions of these complexes are well conserved in eukaryotes, but the extreme expansion of the protein subunit families in plants suggests that plant cells require more regulatory specialization when compared with other eukaryotes. The retromer is associated with retrograde sorting and trafficking of protein cargo back towards the TGN and vacuole in plants, while in animals, there is new evidence that the VPS26C ortholog is associated with recycling or \'retrieving\' proteins back to the PM from the endosomes. The human VPS26C was shown to rescue vps26c mutant phenotypes in Arabidopsis thaliana, suggesting that the retriever function could be conserved in plants. This switch from retromer to retriever function may be associated with core complexes that include the VPS26C subunit in plants, similar to what has been suggested in other eukaryotic systems. We review what is known about retromer function in light of recent findings on functional diversity and specialization of the retromer complex in plants.

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Kinesins, the microtubule-dependent mechanochemical enzymes, power a variety of intracellular movements. Regulation of Kinesin activity and Kinesin-Cargo interactions determine the direction, timing and flux of various intracellular transports. This review examines how phosphorylation of Kinesin subunits and adaptors influence the traffic driven by Kinesin-1, -2, and -3 family motors. Each family of Kinesins are phosphorylated by a partially overlapping set of serine/threonine kinases, and each event produces a unique outcome. For example, phosphorylation of the motor domain inhibits motility, and that of the stalk and tail domains induces cargo loading and unloading effects according to the residue and context. Also, the association of accessory subunits with cargo and adaptor proteins with the motor, respectively, is disrupted by phosphorylation. In some instances, phosphorylation by the same kinase on different Kinesins elicited opposite outcomes. We discuss how this diverse range of effects could manage the logistics of Kinesin-dependent, long-range intracellular transport.

Batten disease is unique among lysosomal storage disorders for the early and profound manifestation in the central nervous system, but little is known regarding potential neuron-specific roles for the disease-associated proteins. We demonstrate substantial overlap in the protein interactomes of three transmembrane Batten proteins (CLN3, CLN6, and CLN8), and that their absence leads to synaptic depletion of key partners (i.e., SNAREs and tethers) and altered synaptic SNARE complexing in vivo, demonstrating a novel shared etiology.

COPII coated vesicles that bud from the endoplasmic reticulum (ER) normally traffic to the Golgi. However, during starvation, COPII vesicles are redirected to the macroautophagy pathway where they become a membrane source for autophagosomes. Phosphorylation of the coat by the casein kinase 1 (CK1), Hrr25, is a prerequisite for vesicle uncoating and membrane fusion. CK1 family members were initially thought to be constitutively active kinases that are regulated through their subcellular localization. Recent studies, however, have shown that the Rab GTPase Ypt1 binds to and activates Hrr25 (CK1δ in mammals) to spatially regulate its kinase activity. Consistent with a direct role for Hrr25 in macroautophagy, hrr25and ypt1mutants are defective in autophagosome biogenesis. These studies have provided insights into how the itinerary of COPII vesicles is coordinated on two different trafficking pathways.

Plasma membrane and membranous organelles contribute to the physiology of the Eukaryotic cell by participating in vesicle trafficking and the maintenance of ion homeostasis. Exomer is a protein complex that facilitates vesicle transport from the trans-Golgi network to the plasma membrane, and its absence leads to the retention of a set of selected cargoes in this organelle. However, this retention does not explain all phenotypes observed in exomer mutants. The Schizosaccharomyces pombe exomer is composed of Cfr1 and Bch1, and cfr1Δ and bch1Δ were sensitive to high concentrations of potassium salts but not sorbitol, which showed sensitivity to ionic but not osmotic stress. Additionally, the activity of the plasma membrane ATPase was higher in exomer mutants than in the wild-type, pointing to membrane hyperpolarization, which caused an increase in intracellular K+ content and mild sensitivity to Na+, Ca2+, and the aminoglycoside antibiotic hygromycin B. Moreover, in response to K+ shock, the intracellular Ca2+ level of cfr1Δ cells increased significantly more than in the wild-type, likely due to the larger Ca2+ spikes in the mutant. Microscopy analyses showed a defective endosomal morphology in the mutants. This was accompanied by an increase in the intracellular pools of the K+ exporting P-type ATPase Cta3 and the plasma membrane Transient Receptor Potential (TRP)-like Ca2+ channel Pkd2, which were partially diverted from the trans-Golgi network to the prevacuolar endosome. Despite this, most Cta3 and Pkd2 were delivered to the plasma membrane at the cell growing sites, showing that their transport from the trans-Golgi network to the cell surface occurred in the absence of exomer. Nevertheless, shortly after gene expression in the presence of KCl, the polarized distribution of Cta3 and Pkd2 in the plasma membrane was disturbed in the mutants. Finally, the use of fluorescent probes suggested that the distribution and dynamics of association of some lipids to the plasma membrane in the presence of KCl were altered in the mutants. Thus, exomer participation in the response to K+ stress was multifaceted. These results supported the notion that exomer plays a general role in protein sorting at the trans-Golgi network and in polarized secretion, which is not always related to a function as a selective cargo adaptor.