Browsing by Author "Kumar, Vimlesh"

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    Modulation of fungal virulence through CRZ1 regulated F-BAR-dependent actin remodeling and endocytosis in chickpea infecting phytopathogen Ascochyta rabiei
    (PLOS, 2021) Sinha, Manisha; Shree, Ankita; Singh, Kunal; Kumar, Kamal; Singh, Shreenivas Kumar; Kumar, Vimlesh; Verma, Praveen K.
    Polarized hyphal growth of filamentous pathogenic fungi is an essential event for host penetration and colonization. The long-range early endosomal trafficking during hyphal growth is crucial for nutrient uptake, sensing of host-specific cues, and regulation of effector production. Bin1/Amphiphysin/Rvs167 (BAR) domain-containing proteins mediate fundamental cellular processes, including membrane remodeling and endocytosis. Here, we identified a F-BAR domain protein (ArF-BAR) in the necrotrophic fungus Ascochyta rabiei and demonstrate its involvement in endosome-dependent fungal virulence on the host plant Cicer arietinum. We show that ArF-BAR regulates endocytosis at the hyphal tip, localizes to the early endosomes, and is involved in actin dynamics. Functional studies involving gene knockout and complementation experiments reveal that ArF-BAR is necessary for virulence. The loss-of-function of ArF-BAR gene results in delayed formation of apical septum in fungal cells near growing hyphal tip that is crucial for host penetration, and impaired secretion of a candidate effector having secretory signal peptide for translocation across the endoplasmic reticulum membrane. The mRNA transcripts of ArF-BAR were induced in response to oxidative stress and infection. We also show that ArF-BAR is able to tubulate synthetic liposomes, suggesting the functional role of F-BAR domain in membrane tubule formation in vivo. Further, our studies identified a stress-induced transcription factor, ArCRZ1 (Calcineurin-responsive zinc finger 1), as key transcriptional regulator of ArF-BAR expression. We propose a model in which ArCRZ1 functions upstream of ArF-BAR to regulate A. rabiei virulence through a mechanism that involves endocytosis, effector secretion, and actin cytoskeleton regulation.
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    Regulation of neuromuscular junction organization by Rab2 and its effector ICA69 in Drosophila
    (The Company of Biologists, 2017) Mallik, Bhagaban; Dwivedi, Manish Kumar; Mushtaq, Zeeshan; Kumari, Manisha; Verma, Praveen K.; Kumar, Vimlesh
    Mechanisms underlying synaptic differentiation, which involves neuronal membrane and cytoskeletal remodeling, are not completely understood. We performed a targeted RNAi-mediated screen of Drosophila BAR-domain proteins and identified islet cell autoantigen 69 kDa (dICA69) as one of the key regulators of morphological differentiation of larval neuromuscular junction (NMJ). We show that Drosophila ICA69 colocalizes with α-Spectrin at the NMJ. The conserved N-BAR domain of dICA69 deforms liposomes in vitro Full length and ICAC but not the N-BAR domain of dICA69 induces filopodia in cultured cells. Consistent with its cytoskeleton regulatory role, dICA69 mutant show reduced α-Spectrin immunoreactivity at the larval NMJ. Manipulating levels of dICA69 or its interactor dPICK1 alters synaptic level of ionotropic glutamate receptors (iGluRs). Moreover, reducing dPICK1 or dRab2 levels phenocopies dICA69 mutation. Interestingly, dRab2 regulates not only synaptic iGluR but also dICA69 levels. Thus, our data suggest that: a) dICA69 regulates NMJ organization through a pathway that involves dPICK1 and dRab2, and b) dRab2 genetically functions upstream of dICA69 and regulates NMJ organization and targeting/retention of iGluRs by regulating dICA69 levels.