@article {152150, title = {Sequence of the SARS-CoV-2 Spike Transmembrane Domain Encodes Conformational Dynamics.}, journal = {J Phys Chem B}, volume = {129}, year = {2025}, month = {2024 Dec 18}, pages = {194-209}, abstract = {

The homotrimeric SARS-CoV-2 spike protein enables viral infection by undergoing a large conformational transition, which facilitates the fusion of the viral envelope with the host cell membrane. The spike protein is anchored to the SARS-CoV-2 envelope by its transmembrane domain (TMD), composed of three TM helices, each contributed by one of the protomers of spike. Although the TMD is known to be important for viral fusion, whether it is a passive anchor of the spike or actively promotes fusion remains unknown. Specifically, it is unclear if the TMD and its dynamics facilitate the prefusion to postfusion conformational transition of the spike. Here, we computationally study the dynamics and self-assembly of the SARS-CoV-2 spike TMD in homogeneous POPC and cholesterol containing membranes. Atomistic simulations of a long TM helix-containing protomer segment show that the membrane-embedded segment bobs, tilts and gains and loses helicity, locally thinning the membrane. Coarse-grained multimerization simulations using representative TM helix structures from the atomistic simulations exhibit diverse trimer populations whose architecture depends on the structure of the TM helix protomer. While a symmetric conformation reflects the symmetry of the resting spike, an asymmetric TMD conformation could promote membrane fusion through the stabilization of a fusion intermediate. Together, our simulations demonstrate that the sequence and length of the SARS-CoV-2 spike TM segment make it inherently dynamic, that trimerization does not abrogate these dynamics and that the various observed TMD conformations may enable viral fusion.

}, issn = {1520-5207}, doi = {10.1021/acs.jpcb.4c05270}, author = {Lall, Sahil and Balaram, Padmanabhan and Mathew, M K and Gosavi, Shachi} } @article {73645, title = {A coherent feed forward loop drives vascular regeneration in damaged aerial organs growing in normal developmental-context.}, journal = {Development}, year = {2020}, month = {2020 Feb 27}, abstract = {

Aerial organs of plants being highly prone to local injuries, require tissue restoration to ensure their survival. However, knowledge of the underlying mechanism is sparse. In this study, we mimicked natural injuries in growing leaf and stem to study the reunion between mechanically disconnected tissues. We show that ()/ () genes, which encodes stem cell promoting factors, are activated and contribute to vascular regeneration in response to these injuries. PLT proteins bind to and activate the CUC2 promoter. Both PLT and CUC2 regulate the transcription of the local auxin biosynthesis gene YUC4 in a coherent feed forward loop, and this process is necessary to drive vascular regeneration. In the absence of this PLT mediated regeneration response, leaf ground tissue cells can neither acquire early vascular identity marker ATHB8, nor properly polarize auxin transporters to specify new venation paths. The PLT-CUC2 module is required for vascular regeneration, but is dispensable for midvein formation in leaf. We reveal the mechanisms of vascular regeneration in plants and distinguishes the wound repair ability of the tissue from its formation during normal development.

}, issn = {1477-9129}, doi = {10.1242/dev.185710}, author = {Radhakrishnan, Dhanya and Shanmukhan, Anju Pallipurath and Kareem, Abdul and Aiyaz, Mohammed and Varapparambathu, Vijina and Toms, Ashna and Kerstens, Merijn and Valsakumar, Devisree and Landge, Amit N and Shaji, Anil and Mathew, M K and Sawchuk, Megan G and Scarpella, Enrico and Krizek, Beth A and Efroni, Idan and M{\"a}h{\"o}nen, Ari Pekka and Willemsen, Viola and Scheres, Ben and Prasad, Kalika} } @article {84574, title = {Role of RAB5 GEF in maintaining potassium levels under sodium chloride stress.}, journal = {Plant Direct}, volume = {4}, year = {2020}, month = {2020 Oct}, pages = {e00273}, abstract = {

Salt stress is one of the major factors impacting crop productivity worldwide. Through a variety of effector and signaling pathways, plants achieve survival under salinity stress by maintaining high cytosolic potassium/sodium ion (K/Na) ratios, preventing Na cytotoxicity, and retaining osmotic balance. Ras-related protein 5 (Rab5) members are involved in the trafficking of endosomes to the vacuole or plasma membrane (PM). The vacuolar protein sorting- associated protein 9 () encodes the single guanine nucleotide exchange factor (GEF) that activates all three known Rab5 proteins in . Previous work from our group has reported the critical function of for the operation of salt-induced endocytic pathway, as well as the expansion of endomembrane compartments under saline stress conditions. Here we show an additional role of in plant response to salt stress via maintenance of K status of the cell rather than Na homeostasis. Our results show that roots from mutant, subjected to 100\ mM NaCl, display alterations in transcript levels of genes involved in the K homeostasis pathway. Concurrent with the observed sensitivity of mutant under NaCl stress, exposure to low K environments resulted in growth retardation, and reduced rate of endocytosis. Furthermore, mutant displays reduced expression of auxin reporter, Direct Repeat-5 (DR5), and alterations in polarity and abundance of auxin efflux carrier PIN- FORMED2 (PIN2). Imposition of NaCl stress was found to be restrictive to the elongation capacity of cells in the root elongation zone of mutant. Together our results indicate that alterations in K homeostasis and associated cellular changes causing increased cell wall pH, contribute to diminished root growth and compromised survival of mutant under salt stress conditions.

}, issn = {2475-4455}, doi = {10.1002/pld3.273}, author = {Rajagopal, Divya and Mathew, M K} } @article {57787, title = {The Ca2+ Channel CNGC19 Regulates Arabidopsis defense against Spodoptera Herbivory.}, journal = {Plant Cell}, year = {2019}, month = {2019 May 10}, abstract = {

Cellular calcium elevation is an important signal used by plants for recognition and signaling of environmental stress. Perception of the generalist insect, Spodoptera litura, by Arabidopsis thaliana activates cytosolic Ca2+ elevation, which triggers downstream defense. However, not all the Ca2+ channels generating the signal have been identified, nor are their modes of action known. We report on a rapidly activated, leaf vasculature- and plasma membrane-localized, CYCLIC NUCLEOTIDE GATED CHANNEL19 (CNGC19), which activates herbivory-induced Ca2+ flux and plant defense. Loss of CNGC19 function results in decreased herbivory defense. The cngc19 mutant shows aberrant and attenuated intra-vascular Ca2+ fluxes. CNGC19 is a Ca2+ permeable channel, as hyperpolarization of CNGC19-expressing Xenopus oocytes in the presence of both cAMP and Ca2+ results in Ca2+ influx. Breakdown of Ca2+-based defence in cngc19 mutants leads to a decrease in herbivory-induced JA-Ile biosynthesis and expression of JA responsive genes. cngc19 mutants are deficient in aliphatic glucosinolate accumulation and hyperaccumulate its precursor, methionine. CNGC19 modulates aliphatic glucosinolate biosynthesis in tandem with BRANCHED-CHAIN AMINO ACID TRANSAMINASE4 (BCAT4), which is involved in the chain elongation pathway of Met-derived glucosinolates. Furthermore, CNGC19 interacts with herbivory-induced CALMODULIN2 (CaM2) in planta. Together, our work reveals a key mechanistic role for the Ca2+ channel CNGC19 in the recognition of herbivory and the activation of defense signaling.

}, issn = {1532-298X}, doi = {10.1105/tpc.19.00057}, author = {Meena, Mukesh Kumar and Prajapati, Ramgopal and Krishna, Deepthi and Divakaran, Keerthi and Pandey, Yogesh and Reichelt, Michael and Mathew, M K and Boland, Wilhelm and Mith{\"o}fer, Axel and Vadassery, Jyothilakshmi} } @article {28565, title = {Fluid flow modulates electrical activity in cardiac hERG potassium channels.}, journal = {J Biol Chem}, year = {2018}, month = {2018 Jan 05}, abstract = {

Fluid movement within the heart generates substantial shear forces, but the effect of this mechanical stress on the electrical activity of the human heart has not been examined. The fast component of the delayed rectifier potassium currents responsible for repolarization of the cardiac action potential, Ikr, is encoded by the hERG channel. Here, we exposed hERG1a channel-expressing HEK293T cells to laminar shear stress (LSS) and observed that this mechanical stress increased the whole-cell current by 30\%-40\%. LSS shifted the voltage dependence of steady-state activation of the hERG\  channel to the hyperpolarizing direction, accelerated the time course of activation and recovery from inactivation, slowed down deactivation, and shifted the steady-state inactivation to the positive direction, all of which favored the hERG open state. In contrast, the time course of inactivation was faster, favoring the closed state. Using specific inhibitors of focal adhesion kinase (FAK), a regulator of mechano-transduction via the integrin pathway, we also found that the LSS-induced modulation of the whole-cell current depended on the integrin pathway. The hERG1b channel variant, which lacks the Per-Arnt-Sim (PAS) domain, and long QT syndrome-associated variants having point mutations in the PAS domain were unaffected by LSS, suggesting that the PAS domain in hERG1a channel may be involved in sensing mechanical shear stress. We conclude that a mechano-electric feedback pathway modulates hERG channel activity through the integrin pathway, indicating that mechanical forces in the heart influence its electrical activity.

}, issn = {1083-351X}, doi = {10.1074/jbc.RA117.000432}, author = {Roy, Samrat and Mathew, M K} } @article {31050, title = {Mechanism of aggregation and membrane interactions of mammalian prion protein.}, journal = {Biochim Biophys Acta}, year = {2018}, month = {2018 Mar 04}, abstract = {

The cellular prion protein (PrP), which is present ubiquitously in all mammalian neurons, is normally found to be linked to the cell membrane through a glycosylphosphatidylinositol (GPI) anchor. The conformational conversion of PrPinto misfolded and aggregated forms is associated with transmissible neurodegenerative diseases known as prion diseases. The importance of different misfolded conformations in prion diseases, and the mechanism by which prion aggregates induce neurotoxicity remain poorly understood. Multiple studies have been shown that the toxicity of misfolded prion protein is directly correlated with its ability to interact with and perturb membranes. This review describes the current progress toward understanding prion protein misfolding and aggregation, as well as the interaction of prion protein aggregates with lipid membrane.

}, issn = {0006-3002}, doi = {10.1016/j.bbamem.2018.02.031}, author = {Sabareesan, Ambadi Thody and Mathew, M K and Udgaonkar, Jayant B} } @article {3540, title = {The Pathogenic A116V Mutation Enhances Ion-Selective Channel Formation by Prion Protein in Membranes.}, journal = {Biophys J}, volume = {110}, year = {2016}, month = {2016 Apr 26}, pages = {1766-76}, abstract = {

Prion diseases are a group of fatal neurodegenerative disorders that afflict mammals. Misfolded and aggregated forms of the prion protein (PrP(Sc)) have been associated with many prion diseases. A transmembrane form of PrP favored by the pathogenic mutation A116V is associated with Gerstmann-Str{\"a}ussler-Scheinker syndrome, but no accumulation of PrP(Sc) is detected. However, the role of the transmembrane form of PrP in pathological processes leading to neuronal death remains unclear. This study reports that the full-length mouse PrP (moPrP) significantly increases the permeability of living cells to K(+), and forms K(+)- and Ca(2+)-selective channels in lipid membranes. Importantly, the pathogenic mutation A116V greatly increases the channel-forming capability of moPrP. The channels thus formed are impermeable to sodium and chloride ions, and are blocked by blockers of voltage-gated ion channels. Hydrogen-deuterium exchange studies coupled with mass spectrometry (HDX-MS) show that upon interaction with lipid, the central hydrophobic region (109-132) of the protein is protected against exchange, making it a good candidate for inserting into the membrane and lining the channel. HDX-MS also shows a dramatic increase in the protein-lipid stoichiometry for A116V moPrP, providing a rationale for its increased channel-forming capability. The results suggest that ion channel formation may be a possible mechanism of PrP-mediated neurodegeneration by the transmembrane forms of PrP.

}, issn = {1542-0086}, doi = {10.1016/j.bpj.2016.03.017}, author = {Sabareesan, Ambadi Thody and Singh, Jogender and Roy, Samrat and Udgaonkar, Jayant B and Mathew, M K} } @article {11658, title = {Regulation of VDAC trafficking modulates cell death.}, journal = {Cell Death Discov}, volume = {2}, year = {2016}, month = {2016}, pages = {16085}, abstract = {

The voltage-dependent anion channel (VDAC) and mitochondria-associated hexokinase (HxK) have crucial roles in both cell survival and death. Both the individual abundances and their ratio seem to influence the balance of survival and death and are thus critical in scenarios, such as neurodegeneration and cancer. Elevated levels of both VDAC and HxK have been reported in cancerous cells. Physical interaction is surmised and specific residues or regions involved have been identified, but details of the interaction and the mechanism by which it modulates survival are yet to be elucidated. We and others have shown that heterologous expression of VDAC can induce cell death, which can be mitigated by concomitant overexpression of HxK. We have also observed that upon overexpression, fluorescently tagged VDAC is distributed between the cytosol and mitochondria. In this study, we show that cell death ensues only when the protein, which is synthesized on cytoplasmic ribosomes, migrates to the mitochondrion. Further, coexpression of rat HxK II (rHxKII) can delay the translocation of human VDAC1 (hVDAC1) protein to mitochondria and thereby inhibit VDAC-induced cell death. Variation in the level of HxK protein as seen endogenously in different cell lines, or as experimentally manipulated by silencing and overexpression, can lead to differential VDAC translocation kinetics and related cell death. The N-terminal region of HxK and the Glu73 residue of hVDAC1, which have previously been implicated in a physical interaction, are required for cytosolic retention of VDAC. Finally, we show that, in otherwise unperturbed cells in culture, there is a small but significant amount of soluble VDAC in the cytosol present in a complex with HxK. This complex could well determine how a cell is poised with respect to incoming thanatopic signals, thereby tilting the survival/death balance in pharmacologically interesting situations, such as neurodegeneration and cancer.

}, doi = {10.1038/cddiscovery.2016.85}, author = {Dubey, Ashvini K and Godbole, Ashwini and Mathew, M K} } @article {871, title = {Expression and purification of OsVDAC4.}, journal = {Methods Enzymol}, volume = {556}, year = {2015}, month = {2015}, pages = {51-75}, abstract = {

The voltage-dependent anion channel (VDAC), a major component of the mitochondrial outer membrane, has emerged as an important player in cell function, survival, and death signaling. VDAC function is modulated by its interaction with proteins such as hexokinase, adenine nucleotide translocator, and apoptotic proteins like Bax. Monitoring the activity of VDAC and its modulation in the complex cellular milieu is fraught with complications. Minimizing the number of components in the study is one approach to teasing apart various aspects of its function. In this chapter, we have described detailed protocols for the purification of a rice VDAC isoform, OsVDAC4 after overexpression in a bacterial system. The protein is solubilized with LDAO and then reconstituted into liposomes or planar bilayers to verify its competence to fold into a functionally active form.

}, issn = {1557-7988}, doi = {10.1016/bs.mie.2014.12.019}, author = {Godbole, Ashwini and Dubey, Ashvini K and Mitra, Rohan and Reddy, Palakolanu S and Mathew, M K} } @article {1042, title = {Functional characterization of a transition metal ion transporter, OsZIP6 from rice (Oryza sativa L.).}, journal = {Plant Physiol Biochem}, volume = {97}, year = {2015}, month = {2015 Dec}, pages = {165-74}, abstract = {

Micronutrients are important for the growth and development of plants, which deploy families of transporters for their uptake and distribution. We have functionally characterized a novel transition metal ion transporter from rice, OsZIP6 (Oryza sativa zinc regulated transporter, iron regulated transporter-like protein 6). The transporter was found to be transcriptionally activated in shoot and root tissues in response to deficiency in Fe(2+), Zn(2+) and Mn(2+). OsZIP6 was expressed in Xenopus laevis oocytes, where currents were observed on addition of Co(2+), Fe(2+) and Cd(2+) but not Zn(2+), Mn(2+) and Ni(2+). This substrate range for OsZIP6, identified using two-electrode voltage clamp electrophysiology was confirmed by atomic absorption spectroscopy. Ion transport by OsZIP6 was found to be pH dependent and enhanced transport was observed at acidic pH. Radioisotope uptake suggested that Co(2+) competitively inhibits Fe(2+) uptake by OsZIP6. Identification and characterization of ZIP family members from crop plants will contribute to an understanding of nutrient mineral homeostasis in these plants.

}, issn = {1873-2690}, doi = {10.1016/j.plaphy.2015.10.005}, author = {P G, Kavitha and Kuruvilla, Sam and Mathew, M K} } @article {882, title = {Salt-induced remodeling of spatially restricted clathrin-independent endocytic pathways in Arabidopsis root.}, journal = {Plant Cell}, volume = {27}, year = {2015}, month = {2015 Apr}, pages = {1297-315}, abstract = {

Endocytosis is a ubiquitous cellular process that is characterized well in animal cells in culture but poorly across intact, functioning tissue. Here, we analyze endocytosis throughout the Arabidopsis thaliana root using three classes of probes: a lipophilic dye, tagged transmembrane proteins, and a lipid-anchored protein. We observe a stratified distribution of endocytic processes. A clathrin-dependent endocytic pathway that internalizes transmembrane proteins functions in all cell layers, while a sterol-dependent, clathrin-independent pathway that takes up lipid and lipid-anchored proteins but not transmembrane proteins is restricted to the epidermal layer. Saline stress induces a third pathway that is clathrin-independent, nondiscriminatory in its choice of cargo, and operates across all layers of the root. Concomitantly, small acidic compartments in inner cell layers expand to form larger vacuole-like structures. Plants lacking function of the Rab-GEF (guanine nucleotide exchange factor) VPS9a (vacuolar protein sorting 9A) neither induce the third endocytic pathway nor expand the vacuolar system in response to salt stress. The plants are also hypersensitive to salt. Thus, saline stress reconfigures clathrin-independent endocytosis and remodels endomembrane systems, forming large vacuoles in the inner cell layers, both processes correlated by the requirement of VPS9a activity.

}, keywords = {Arabidopsis, Biological Transport, Clathrin, Endocytosis, Plant Roots, Signal Transduction, Sodium Chloride}, issn = {1532-298X}, doi = {10.1105/tpc.15.00154}, author = {Baral, Anirban and Irani, Niloufer G and Fujimoto, Masaru and Nakano, Akihiko and Mayor, Satyajit and Mathew, M K} } @article {1018, title = {Vesicular trafficking and salinity responses in plants.}, journal = {IUBMB Life}, volume = {67}, year = {2015}, month = {2015 Sep}, pages = {677-86}, abstract = {

Research spanning three decades has demonstrated that vesicles pinch off from the plasma membrane and traffic through the cytoplasm of plant cells, much as previously reported in animal cells. Although the well-conserved clathrin-mediated mechanism of endocytosis has been well characterized, relatively little is known about clathrin-independent pathways in plants. Modulation of endocytosis by both physical stimuli and chemical ligands has been reported in plants. Here, we review the effect of salinity-one of the most deleterious environmental assaults-on endocytosis and intracellular trafficking. {\textcopyright} 2015 IUBMB Life, 67(9):677-686, 2015.

}, issn = {1521-6551}, doi = {10.1002/iub.1425}, author = {Baral, Anirban and Shruthi, K S and Mathew, M K} }