Legionella effectors SidC/SdcA ubiquitinate multiple small GTPases and SNARE proteins to promote phagosomal maturation.

Protein ubiquitination is one of the most important posttranslational modifications (PTMs) in eukaryotes and is involved in the regulation of almost all cellular signaling pathways. The intracellular bacterial pathogen Legionella pneumophila translocates at least 26 effectors to hijack host ubiquitination signaling via distinct mechanisms. Among these effectors, SidC/SdcA are novel E3 ubiquitin ligases with the adoption of a Cys-His-Asp catalytic triad. SidC/SdcA are critical for the recruitment of endoplasmic reticulum (ER)-derived vesicles to the Legionella-containing vacuole (LCV). However, the ubiquitination targets of SidC/SdcA are largely unknown, which restricts our understanding of the mechanisms used by these effectors to hijack the vesicle trafficking pathway. Here, we demonstrated that multiple Rab small GTPases and target soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARE) proteins are bona fide ubiquitination substrates of SidC/SdcA. SidC/SdcA-mediated ubiquitination of syntaxin 3 and syntaxin 4 promotes their unconventional pairing with the vesicle-SNARE protein Sec22b, thereby contributing to the membrane fusion of ER-derived vesicles with the phagosome. In addition, our data reveal that ubiquitination of Rab7 by SidC/SdcA is critical for its association with the LCV membrane. Rab7 ubiquitination could impair its binding with the downstream effector Rab-interacting lysosomal protein (RILP), which partially explains why LCVs avoid fusion with lysosomes despite the acquisition of Rab7. Taken together, our study reveals the biological mechanisms employed by SidC/SdcA to promote the maturation of the LCVs.

Coxiella burnetii-containing vacuoles interact with host recycling endosomal proteins Rab11a and Rab35 for vacuolar expansion and bacterial growth.

Introduction: Coxiella burnetii is a gram-negative obligate intracellular bacterium and a zoonotic pathogen that causes human Q fever. The lack of effective antibiotics and a licensed vaccine for Coxiella in the U.S. warrants further research into Coxiella pathogenesis. Within the host cells, Coxiella replicates in an acidic phagolysosome-like vacuole termed Coxiella-containing vacuole (CCV). Previously, we have shown that the CCV pH is critical for Coxiella survival and that the Coxiella Type 4B secretion system regulates CCV pH by inhibiting the host endosomal maturation pathway. However, the trafficking pattern of the 'immature' endosomes in Coxiella- infected cells remained unclear. Methods: We transfected HeLa cells with GFP-tagged Rab proteins and subsequently infected them with mCherry-Coxiella to visualize Rab protein localization. Infected cells were immunostained with anti-Rab antibodies to confirm the Rab localization to the CCV, to quantitate Rab11a and Rab35- positive CCVs, and to quantitate total recycling endosome content of infected cells. A dual-hit siRNA mediated knockdown combined with either immunofluorescent assay or an agarose-based colony-forming unit assay were used to measure the effects of Rab11a and Rab35 knockdown on CCV area and Coxiella intracellular growth. Results: The CCV localization screen with host Rab proteins revealed that recycling endosome-associated proteins Rab11a and Rab35 localize to the CCV during infection, suggesting that CCV interacts with host recycling endosomes during maturation. Interestingly, only a subset of CCVs were Rab11a or Rab35-positive at any given time point. Quantitation of Rab11a/Rab35-positive CCVs revealed that while Rab11a interacts with the CCV more at 3 dpi, Rab35 is significantly more prevalent at CCVs at 6 dpi, suggesting that the CCV preferentially interacts with Rab11a and Rab35 depending on the stage of infection. Furthermore, we observed a significant increase in Rab11a and Rab35 fluorescent intensity in Coxiella-infected cells compared to mock, suggesting that Coxiella increases the recycling endosome content in infected cells. Finally, siRNA-mediated knockdown of Rab11a and Rab35 resulted in significantly smaller CCVs and reduced Coxiella intracellular growth, suggesting that recycling endosomal Rab proteins are essential for CCV expansion and bacterial multiplication. Discussion: Our data, for the first time, show that the CCV dynamically interacts with host recycling endosomes for Coxiella intracellular survival and potentially uncovers novel host cell factors essential for Coxiella pathogenesis.

Initiation of acute pancreatitis in mice is independent of fusion between lysosomes and zymogen granules.

The co-localization of the lysosomal protease cathepsin B (CTSB) and the digestive zymogen trypsinogen is a prerequisite for the initiation of acute pancreatitis. However, the exact molecular mechanisms of co-localization are not fully understood. In this study, we investigated the role of lysosomes in the onset of acute pancreatitis by using two different experimental approaches. Using an acinar cell-specific genetic deletion of the ras-related protein Rab7, important for intracellular vesicle trafficking and fusion, we analyzed the subcellular distribution of lysosomal enzymes and the severity of pancreatitis in vivo and ex vivo. Lysosomal permeabilization was performed by the lysosomotropic agent Glycyl-L-phenylalanine 2-naphthylamide (GPN). Acinar cell-specific deletion of Rab7 increased endogenous CTSB activity and despite the lack of re-distribution of CTSB from lysosomes to the secretory vesicles, the activation of CTSB localized in the zymogen compartment still took place leading to trypsinogen activation and pancreatic injury. Disease severity was comparable to controls during the early phase but more severe at later time points. Similarly, GPN did not prevent CTSB activation inside the secretory compartment upon caerulein stimulation, while lysosomal CTSB shifted to the cytosol. Intracellular trypsinogen activation was maintained leading to acute pancreatitis similar to controls. Our results indicate that initiation of acute pancreatitis seems to be independent of the presence of lysosomes and that fusion of lysosomes and zymogen granules is dispensable for the disease onset. Intact lysosomes rather appear to have protective effects at later disease stages.

Molecular mechanism for recognition of the cargo adapter Rab6GTP by the dynein adapter BicD2.

Rab6 is a key modulator of protein secretion. The dynein adapter Bicaudal D2 (BicD2) recruits the motors cytoplasmic dynein and kinesin-1 to Rab6GTP-positive vesicles for transport; however, it is unknown how BicD2 recognizes Rab6. Here, we establish a structural model for recognition of Rab6GTP by BicD2, using structure prediction and mutagenesis. The binding site of BicD2 spans two regions of Rab6 that undergo structural changes upon the transition from the GDP- to GTP-bound state, and several hydrophobic interface residues are rearranged, explaining the increased affinity of the active GTP-bound state. Mutations of Rab6GTP that abolish binding to BicD2 also result in reduced co-migration of Rab6GTP/BicD2 in cells, validating our model. These mutations also severely diminished the motility of Rab6-positive vesicles in cells, highlighting the importance of the Rab6GTP/BicD2 interaction for overall motility of the multi-motor complex that contains both kinesin-1 and dynein. Our results provide insights into trafficking of secretory and Golgi-derived vesicles and will help devise therapies for diseases caused by BicD2 mutations, which selectively affect the affinity to Rab6 and other cargoes.

RUFY4 deletion prevents pathological bone loss by blocking endo-lysosomal trafficking of osteoclasts.

Mature osteoclasts degrade bone matrix by exocytosis of active proteases from secretory lysosomes through a ruffled border. However, the molecular mechanisms underlying lysosomal trafficking and secretion in osteoclasts remain largely unknown. Here, we show with GeneChip analysis that RUN and FYVE domain-containing protein 4 (RUFY4) is strongly upregulated during osteoclastogenesis. Mice lacking Rufy4 exhibited a high trabecular bone mass phenotype with abnormalities in osteoclast function in vivo. Furthermore, deleting Rufy4 did not affect osteoclast differentiation, but inhibited bone-resorbing activity due to disruption in the acidic maturation of secondary lysosomes, their trafficking to the membrane, and their secretion of cathepsin K into the extracellular space. Mechanistically, RUFY4 promotes late endosome-lysosome fusion by acting as an adaptor protein between Rab7 on late endosomes and LAMP2 on primary lysosomes. Consequently, Rufy4-deficient mice were highly protected from lipopolysaccharide- and ovariectomy-induced bone loss. Thus, RUFY4 plays as a new regulator in osteoclast activity by mediating endo-lysosomal trafficking and have a potential to be specific target for therapies against bone-loss diseases such as osteoporosis.

TBC9, an essential TBC-domain protein, regulates early vesicular transport and IMC formation in Toxoplasma gondii.

Apicomplexan parasites harbor a complex endomembrane system as well as unique secretory organelles. These complex cellular structures require an elaborate vesicle trafficking system, which includes Rab GTPases and their regulators, to assure the biogenesis and secretory of the organelles. Here we exploit the model apicomplexan organism Toxoplasma gondii that encodes a family of Rab GTPase Activating Proteins, TBC (Tre-2/Bub2/Cdc16) domain-containing proteins. Functional profiling of these proteins in tachyzoites reveals that TBC9 is the only essential regulator, which is localized to the endoplasmic reticulum (ER) in T. gondii strains. Detailed analyses demonstrate that TBC9 is required for normal distribution of proteins targeting to the ER, and the Golgi apparatus in the parasite, as well as for the normal formation of daughter inner membrane complexes (IMCs). Pull-down assays show a strong protein interaction between TBC9 and specific Rab GTPases (Rab11A, Rab11B, and Rab2), supporting the role of TBC9 in daughter IMC formation and early vesicular transport. Thus, this study identifies the only essential TBC domain-containing protein TBC9 that regulates early vesicular transport and IMC formation in T. gondii and potentially in closely related protists.

The vacuolar fusion regulated by HOPS complex promotes hyphal initiation and penetration in Candida albicans.

The transition between yeast and hyphae is crucial for regulating the commensalism and pathogenicity in Candida albicans. The mechanisms that affect the invasion of hyphae in solid media, whose deficiency is more related to the pathogenicity of C. albicans, have not been elucidated. Here, we found that the disruption of VAM6 or VPS41 which are components of the homotypic vacuolar fusion and protein sorting (HOPS) complex, or the Rab GTPase YPT72, all responsible for vacuole fusion, led to defects in hyphal growth in both liquid and solid media, but more pronounced on solid agar. The phenotypes of vac8Δ/Δ and GTR1OE-vam6Δ/Δ mutants indicated that these deficiencies are mainly caused by the reduced mechanical forces that drive agar and organs penetration, and confirmed that large vacuoles are required for hyphal mechanical penetration. In summary, our study revealed that large vacuoles generated by vacuolar fusion support hyphal penetration and provided a perspective to refocus attention on the role of solid agar in evaluating C. albicans invasion.

Pseudorabies virus hijacks the Rab6 protein to promote viral assembly and egress.

Pseudorabies virus (PRV) is recognized as the aetiological agent responsible for Aujeszky's disease, or pseudorabies, in swine populations. Rab6, a member of the small GTPase family, is implicated in various membrane trafficking processes, particularly exocytosis regulation. Its involvement in PRV infection, however, has not been documented previously. In our study, we observed a significant increase in the Rab6 mRNA and protein levels in both PK-15 porcine kidney epithelial cells and porcine alveolar macrophages, as well as in the lungs and spleens of mice infected with PRV. The overexpression of wild-type Rab6 and its GTP-bound mutant facilitated PRV proliferation, whereas the GDP-bound mutant form of Rab6 had no effect on viral propagation. These findings indicated that the GTPase activity of Rab6 was crucial for the successful spread of PRV. Further investigations revealed that the reduction in Rab6 levels through knockdown significantly hampered PRV proliferation and disrupted virus assembly and egress. At the molecular level, Rab6 was found to interact with the PRV glycoproteins gB and gE, both of which are essential for viral assembly and egress. Our results collectively suggest that PRV exploits Rab6 to expedite its assembly and egress and identify Rab6 as a promising novel target for therapeutic treatment for PRV infection.

Legionella pneumophila exploits the endo-lysosomal network for phagosome biogenesis by co-opting SUMOylated Rab7.

Legionella pneumophila strains harboring wild-type rpsL such as Lp02rpsLWT cannot replicate in mouse bone marrow-derived macrophages (BMDMs) due to induction of extensive lysosome damage and apoptosis. The bacterial factor directly responsible for inducing such cell death and the host factor involved in initiating the signaling cascade that leads to lysosome damage remain unknown. Similarly, host factors that may alleviate cell death induced by these bacterial strains have not yet been investigated. Using a genome-wide CRISPR/Cas9 screening, we identified Hmg20a and Nol9 as host factors important for restricting strain Lp02rpsLWT in BMDMs. Depletion of Hmg20a protects macrophages from infection-induced lysosomal damage and apoptosis, allowing productive bacterial replication. The restriction imposed by Hmg20a was mediated by repressing the expression of several endo-lysosomal proteins, including the small GTPase Rab7. We found that SUMOylated Rab7 is recruited to the bacterial phagosome via SulF, a Dot/Icm effector that harbors a SUMO-interacting motif (SIM). Moreover, overexpression of Rab7 rescues intracellular growth of strain Lp02rpsLWT in BMDMs. Our results establish that L. pneumophila exploits the lysosomal network for the biogenesis of its phagosome in BMDMs.

Function and regulation of Rab GTPases in cancers.

The Rab small GTPases are characterized by the distinct intracellular localization and modulate various endocytic, transcytic and exocytic transport pathways. Rab proteins function as scaffolds that connect signaling pathways and intracellular membrane trafficking processes through the recruitment of effectors, such as tethering factors, phosphatases, motors and kinases. In different cancers, Rabs play as either an onco-protein or a tumor suppressor role, highly dependending on the context. The molecular mechanistic research has revealed that Rab proteins are involved in cancer progression through influences on migration, invasion, metabolism, exosome secretion, autophagy, and drug resistance of cancer cells. Therefore, targeting Rab GTPases to recover the dysregulated vesicle transport systems may provide potential strategy to restrain cancer progression. In this review, we discuss the regulation of Rab protein level and activity in modulating pathways involved in tumor progression, and propose that Rab proteins may serve as a prognostic factor in different cancers.

Multi-tiered actions of Legionella effectors to modulate host Rab10 dynamics.

Rab GTPases are representative targets of manipulation by intracellular bacterial pathogens for hijacking membrane trafficking. Legionella pneumophila recruits many Rab GTPases to its vacuole and exploits their activities. Here, we found that infection-associated regulation of Rab10 dynamics involves ubiquitin signaling cascades mediated by the SidE and SidC families of Legionella ubiquitin ligases. Phosphoribosyl-ubiquitination of Rab10 catalyzed by the SidE ligases is crucial for its recruitment to the bacterial vacuole. SdcB, the previously uncharacterized SidC-family effector, resides on the vacuole and contributes to retention of Rab10 at the late stages of infection. We further identified MavC as a negative regulator of SdcB. By the transglutaminase activity, MavC crosslinks ubiquitin to SdcB and suppresses its function, resulting in elimination of Rab10 from the vacuole. These results demonstrate that the orchestrated actions of many L. pneumophila effectors fine-tune the dynamics of Rab10 during infection.

Rab30 facilitates lipid homeostasis during fasting.

To facilitate inter-tissue communication and the exchange of proteins, lipoproteins, and metabolites with the circulation, hepatocytes have an intricate and efficient intracellular trafficking system regulated by small Rab GTPases. Here, we show that Rab30 is induced in the mouse liver by fasting, which is amplified in liver-specific carnitine palmitoyltransferase 2 knockout mice (Cpt2L-/-) lacking the ability to oxidize fatty acids, in a Pparα-dependent manner. Live-cell super-resolution imaging and in vivo proximity labeling demonstrates that Rab30-marked vesicles are highly dynamic and interact with proteins throughout the secretory pathway. Rab30 whole-body, liver-specific, and Rab30; Cpt2 liver-specific double knockout (DKO) mice are viable with intact Golgi ultrastructure, although Rab30 deficiency in DKO mice suppresses the serum dyslipidemia observed in Cpt2L-/- mice. Corresponding with decreased serum triglyceride and cholesterol levels, DKO mice exhibit decreased circulating but not hepatic ApoA4 protein, indicative of a trafficking defect. Together, these data suggest a role for Rab30 in the selective sorting of lipoproteins to influence hepatocyte and circulating triglyceride levels, particularly during times of excessive lipid burden.

Biogenic selenium nanoparticles alleviate intestinal barrier injury in mice through TBC1D15/Fis1/Rab7 pathway.

Intestinal diseases often stem from a compromised intestinal barrier. This barrier relies on a functional epithelium and proper turnover of intestinal cells, supported by mitochondrial health. Mitochondria and lysosomes play key roles in cellular balance. Our previous researches indicate that biogenic selenium nanoparticles (SeNPs) can alleviate intestinal epithelial barrier damage by enhancing mitochondria-lysosome crosstalk, though the detailed mechanism is unclear. This study aimed to investigate the role of mitochondria-lysosome crosstalk in the protective effect of SeNPs on intestinal barrier function in mice exposed to lipopolysaccharide (LPS). The results showed that LPS exposure increased intestinal permeability in mice, leding to structural and functional damage to mitochondrial and lysosomal. Oral administration of SeNPs significantly upregulated the expression levels of TBC1D15 and Fis1, downregulated the expression levels of Rab7, Caspase-3, Cathepsin B, and MCOLN2, effectively alleviated LPS-induced mitochondrial and lysosomal dysfunction and maintained the intestinal barrier integrity in mice. Furthermore, SeNPs notably inhibited mitophagy caused by adenovirus-associated virus (AAV)-mediated RNA interference the expression of TBC1D15 in the intestine of mice, maintained mitochondrial and lysosomal homeostasis, and effectively alleviated intestinal barrier damage. These results suggested that SeNPs can regulate mitochondria-lysosome crosstalk and inhibit its damage by regulating the TBC1D15/Fis1/Rab7- signaling pathway. thereby alleviating intestinal barrier damage. It lays a theoretical foundation for elucidating the mechanism of mitochondria-lysosome crosstalk in regulating intestinal barrier damage and repair, and provides new ideas and new ways to establish safe and efficient nutritional regulation strategies to prevent and treat intestinal diseases caused by inflammation.

IL-33/NF-κB/ST2L/Rab37 positive-feedback loop promotes M2 macrophage to limit chemotherapeutic efficacy in lung cancer.

IL-33 is a danger signal that binds to its receptor ST2L to promote tumor progression. This study identifies the IL-33/ST2L positive-feedback loop and the trafficking of ST2L membrane presentation in macrophages that contribute to lung tumor progression. Mechanistically, IL-33 induces ST2L upregulation by activating NF-κB, which binds to the promoter region of the ST2L gene. Moreover, Rab37, a small GTPase involved in membrane trafficking, mediates ST2L trafficking to the plasma membrane of M2 macrophages. This IL-33/NF-κB/ST2L/Rab37 axis promotes positive-feedback loops that enhance ST2L expression and membrane trafficking in M2 macrophages. Notably, neutralizing antibodies against IL-33 or ST2L block NF-κB activity, suppress M2 macrophage polarization, and synergistically inhibit tumor growth when combined with cisplatin treatment in vitro/vivo. Clinically, Rab37+/ST2L+/CD206+ tumor-infiltrating M2 macrophages correlate with advanced-stage lung cancer patients with poor response to chemotherapy. These findings unveil a positive-feedback mechanism and provide a basis for IL-33/ST2L-targeting therapy for cancer.

Systematic identification of structure-specific protein-protein interactions.

The physical interactome of a protein can be altered upon perturbation, modulating cell physiology and contributing to disease. Identifying interactome differences of normal and disease states of proteins could help understand disease mechanisms, but current methods do not pinpoint structure-specific PPIs and interaction interfaces proteome-wide. We used limited proteolysis-mass spectrometry (LiP-MS) to screen for structure-specific PPIs by probing for protease susceptibility changes of proteins in cellular extracts upon treatment with specific structural states of a protein. We first demonstrated that LiP-MS detects well-characterized PPIs, including antibody-target protein interactions and interactions with membrane proteins, and that it pinpoints interfaces, including epitopes. We then applied the approach to study conformation-specific interactors of the Parkinson's disease hallmark protein alpha-synuclein (aSyn). We identified known interactors of aSyn monomer and amyloid fibrils and provide a resource of novel putative conformation-specific aSyn interactors for validation in further studies. We also used our approach on GDP- and GTP-bound forms of two Rab GTPases, showing detection of differential candidate interactors of conformationally similar proteins. This approach is applicable to screen for structure-specific interactomes of any protein, including posttranslationally modified and unmodified, or metabolite-bound and unbound protein states.

Rab11 maintains the undifferentiated state of adult midgut precursors via DPP pathway.

Asymmetric stem cell divisions play instrumental roles in the maintenance, growth and differentiation of organs. Failure of asymmetric stem cell divisions may result in an array of developmental disorders, including cancer. It is well established that the gene, inscuteable, acts as the upstream component of asymmetric cell divisions. In Drosophila larval midgut, a founder adult midgut precursor (AMP) experiences an asymmetric division to instruct its first daughter to become a peripheral cell that serves as a niche where the AMP and its future daughters can remain undifferentiated. The present study demonstrates that inscuteable expressing stem cells require Rab11, a conserved small Ras-like GTPase, for proper proliferation and differentiation. As insc-GAL4 mediated Rab11RNAi in Drosophila larval and adult midguts show the disruption of the niche microenvironment of adult midgut precursors as well as elevated DPP signalling at the larval stage, which is associated with aberrant over-proliferation and early differentiation of larval AMPs and adult intestinal stem cells. The observed connections between Rab11, larval AMP proliferation, niche establishment, and DPP signalling highlight the potential for Rab11 to serve as a key regulatory factor in maintaining tissue homeostasis and balanced cellular growth.

Loss of RAB25 Cooperates with Oncogenes in the Transformation of Human Mammary Epithelial Cells (HMECs) to Give Rise to Claudin-Low Tumors.

The loss of RAB25 expression-RAS superfamily of GTPase characteristic of numerous breast cancers-corresponds with H-RAS point mutations, particularly in triple-negative breast cancers (TNBC), a subtype associated with a poor prognosis. To address the poorly understood factors dictating the progression of TNBC tumors, we examine the cooperative effects that loss of RAB25 expression in human mammary epithelial cell (HMEC) lines with H-RAS mutations confers in tumorigenesis. HMECs were immortalized by transduction with LXSN CDK4 R24C, a mutant form of cyclin-dependent kinase, followed by transduction with hTERT, a catalytic subunit of the telomerase enzyme. We found that with the loss of RAB25 and overexpression of mutant H-RAS61L, immortal HMECs transformed toward anchorage-independent growth and acquired an increased ability to migrate. Furthermore, cells express low CD24, high CD44, and low claudin levels, indicating stem-like properties upon transformation. Besides, loss of RAB25 and overexpression of H-RAS61L resulted in increased expression of transcription factors Snail and Slug that drive these cells to lose E-cadherin and undergo epithelial-mesenchymal transition (EMT). This study confirms that loss of RAB25 and overexpression of mutant H-RAS can drive HMECs toward a mesenchymal stem-like state. Our findings reveal that RAB25 functions as a tumor suppressor gene, and loss of RAB25 could serve as a novel biomarker of the claudin-low type of TNBC.

Porcine reproductive and respiratory syndrome virus 2 hijacks CMA-mediated lipolysis through upregulation of small GTPase RAB18.

RAB GTPases (RABs) control intracellular membrane trafficking with high precision. In the present study, we carried out a short hairpin RNA (shRNA) screen focused on a library of 62 RABs during infection with porcine reproductive and respiratory syndrome virus 2 (PRRSV-2), a member of the family Arteriviridae. We found that 13 RABs negatively affect the yield of PRRSV-2 progeny virus, whereas 29 RABs have a positive impact on the yield of PRRSV-2 progeny virus. Further analysis revealed that PRRSV-2 infection transcriptionally regulated RAB18 through RIG-I/MAVS-mediated canonical NF-κB activation. Disrupting RAB18 expression led to the accumulation of lipid droplets (LDs), impaired LDs catabolism, and flawed viral replication and assembly. We also discovered that PRRSV-2 co-opts chaperone-mediated autophagy (CMA) for lipolysis via RAB18, as indicated by the enhanced associations between RAB18 and perlipin 2 (PLIN2), CMA-specific lysosomal associated membrane protein 2A (LAMP2A), and heat shock protein family A (Hsp70) member 8 (HSPA8/HSC70) during PRRSV-2 infection. Knockdown of HSPA8 and LAMP2A impacted on the yield of PRRSV-2 progeny virus, implying that the virus utilizes RAB18 to promote CMA-mediated lipolysis. Importantly, we determined that the C-terminal domain (CTD) of HSPA8 could bind to the switch II domain of RAB18, and the CTD of PLIN2 was capable of associating with HSPA8, suggesting that HSPA8 facilitates the interaction between RAB18 and PLIN2 in the CMA process. In summary, our findings elucidate how PRRSV-2 hijacks CMA-mediated lipid metabolism through innate immune activation to enhance the yield of progeny virus, offering novel insights for the development of anti-PRRSV-2 treatments.

The role of Rab GTPase in Plant development and stress.

Small GTPase is a type of crucial regulator in eukaryotes. It acts as a molecular switch by binding with GTP and GDP in cytoplasm, affecting various cellular processes. Small GTPase were divided into five subfamilies based on sequence, structure and function: Ras, Rho, Rab, Arf/Sar and Ran, with Rab being the largest subfamily. Members of the Rab subfamily play an important role in regulating complex vesicle transport and microtubule system activity. Plant cells are composed of various membrane-bound organelles, and vesicle trafficking is fundamental to the existence of plants. At present, the function of some Rab members, such as RabA1a, RabD2b/c and RabF2, has been well characterized in plants. This review summarizes the role of Rab GTPase in regulating plant tip growth, morphogenesis, fruit ripening and stress response, and briefly describes the regulatory mechanisms involved. It provides a reference for further alleviating environmental stress, improving plant resistance and even improving fruit quality.

Post-translational targeting of Rab35 by the effector IcsB of Shigella determines intracellular bacterial niche formation.

Escape from the bacterial-containing vacuole (BCV) is a key step of Shigella host cell invasion. Rab GTPases subverted to in situ-formed macropinosomes in the vicinity of the BCV have been shown to promote its rupture. The involvement of the BCV itself has remained unclear. We demonstrate that Rab35 is non-canonically entrapped at the BCV. Stimulated emission depletion imaging localizes Rab35 directly on the BCV membranes before vacuolar rupture. The bacterial effector IcsB, a lysine Nε-fatty acylase, is a key regulator of Rab35-BCV recruitment, and we show post-translational acylation of Rab35 by IcsB in its polybasic region. While Rab35 and IcsB are dispensable for the first step of BCV breakage, they are needed for the unwrapping of damaged BCV remnants from Shigella. This provides a framework for understanding Shigella invasion implicating re-localization of a Rab GTPase via its bacteria-dependent post-translational modification to support the mechanical unpeeling of the BCV.