Viral spike proteins mutate frequently, but conserved features within these proteins often have functional importance and can inform development of anti-viral therapies which circumvent the effects of viral sequence mutations. Through analysis of large numbers of viral spike protein sequences from several viral families, we found highly (>99%) conserved patterns within their intracellular domains. The patterns generally consist of one or more basic amino acids (arginine or lysine) adjacent to a cysteine, many of which are known to undergo acylation. These patterns were not enriched in cellular proteins in general. Molecular dynamics simulations show direct electrostatic and hydrophobic interactions between these conserved residues in hemagglutinin (HA) from influenza A and B and the phosphoinositide PIP2. Super-resolution microscopy shows nanoscale colocalization of PIP2 and several of the same viral proteins. We propose the hypothesis that these conserved viral spike protein features can interact with phosphoinositides such as PIP2.

The formation of autophagosomes mediating the sequestration of cytoplasmic materials is the central step of autophagy. Several phosphoinositides, which are signaling molecules on the membrane, are involved in autophagy. However, it is not always clear whether these phosphoinositides act directly at the site of autophagosome formation, or indirectly via the regulation of other steps or pathways. To address this question, we used a set of phosphoinositide probes to systematically examine their potential presence on autophagosomal membranes in yeast (Saccharomyces cerevisiae). We verified the specificity of these probes using mutant cells deficient in the production of the corresponding phosphoinositides. We then examined starved yeast cells co-expressing a phosphoinositide probe together with an autophagosomal membrane marker, 2Katushka2S-Atg8. Our data revealed that PtdIns(4,5)P2 and PtdIns(3,5)P2 were mainly present on the plasma membrane and vacuolar membrane, respectively. We observed only occasional co-localization between the PtdIns(4)P probe and Atg8, some of which may represent the transient passage of a PtdIns(4)P-containing structure near the autophagosomal membrane. In contrast, substantial colocalization of the PtdIns(3)P probe with Atg8 was observed. Taken together, our data indicate that only PtdIns(3)P is present in a substantial amount on the autophagosomal membrane. For other phosphoinositides involved in autophagy, either their presence on the autophagosomal membrane is very transient, or they act on other cellular membranes to regulate autophagy.

Septins are cytoskeletal proteins and their interaction with membranes is crucial for their role in various cellular processes. Septins have polybasic regions (PB1 and PB2) which are important for lipid interaction. Earlier, we and others have highlighted the role of the septin C-terminal domain (CTD) to membrane interaction. However, detailed information on residues/group of residues important for such feature is lacking. In this study, we investigate the lipid-binding profile of Schistosoma mansoni Septin10 (SmSEPT10) using PIP strip and Langmuir monolayer adsorption assays. Our findings highlight the CTD as the primary domain responsible for lipid interaction in SmSEPT10, showing binding to phosphatidylinositol phosphates. SmSEPT10 CTD contains a conserved polybasic region (PB3) present in both animals and fungi septins, and a Lys (K367) within its putative amphipathic helix (AH) that we demonstrate as important for lipid binding. PB3 deletion or mutation of this Lys (K367A) strongly impairs lipid interaction. Remarkably, we observe that the AH within a construct lacking the final 43 amino acid residues is insufficient for lipid binding. Furthermore, we investigate the homocomplex formed by SmSEPT10 CTD in solution by cross-linking experiments, CD spectroscopy, SEC-MALS and SEC-SAXS. Taken together, our studies define the lipid-binding region in SmSEPT10 and offer insights into the molecular basis of septin-membrane binding. This information is particularly relevant for less-studied non-human septins, such as SmSEPT10.

PtdIns and its phosphorylated derivatives, the phosphoinositides, are the biochemical components of a major pathway of intracellular signaling in all eukaryotic cells. These lipids are few in terms of cohort of unique positional isomers, and are quantitatively minor species of the bulk cellular lipidome. Nevertheless, phosphoinositides regulate an impressively diverse set of biological processes. It is from that perspective that perturbations in phosphoinositide-dependent signaling pathways are increasingly being recognized as causal foundations of many human diseases - including cancer. Although phosphatidylinositol transfer proteins (PITPs) are not enzymes, these proteins are physiologically significant regulators of phosphoinositide signaling. As such, PITPs are conserved throughout the eukaryotic kingdom. Their biological importance notwithstanding, PITPs remain understudied. Herein, we review current information regarding PITP biology primarily focusing on how derangements in PITP function disrupt key signaling/developmental pathways and are associated with a growing list of pathologies in mammals.

Adaptive immune responses comprise the activation of T cells by peptide antigens that are presented by proteins of the Major Histocompatibility Complex (MHC) on the surface of an antigen-presenting cell. As a consequence of the T cell receptor interacting productively with a certain peptide-MHC complex, a specialized cell-cell junction known as the immunological synapse forms and is accompanied by changes in the spatiotemporal patterning and function of intracellular signaling molecules. Key modifications occurring at the cytoplasmic leaflet of the plasma and internal membranes in activated T cells comprise lipid switches that affect the binding and distribution of proteins within or near the lipid bilayer. Here, we describe two major classes of lipid switches that act at this critical water/membrane interface. Phosphoinositides are derived from phosphatidylinositol, an amphiphilic molecule that contains two fatty acid chains and a phosphate group that bridges the glycerol backbone to the carbohydrate inositol. The inositol ring can be variably (de-)phosphorylated by dedicated kinases and phosphatases, thereby creating phosphoinositide signatures that define the composition and properties of signaling molecules, molecular complexes, or whole organelles. Palmitoylation refers to the reversible attachment of the fatty acid palmitate to a substrate protein\'s cysteine residue. DHHC enzymes, named after the four conserved amino acids in their active site, catalyze this post-translational modification and thereby change the distribution of proteins at, between, and within membranes. T cells utilize these two types of molecular switches to adjust their properties to an activation process that requires changes in motility, transport, secretion, and gene expression.

Myogenesis is a multistep process that requires a spatiotemporal regulation of cell events resulting finally in myoblast fusion into multinucleated myotubes. Most major insights into the mechanisms underlying fusion seem to be conserved from insects to mammals and include the formation of podosome-like protrusions (PLPs) that exert a driving force toward the founder cell. However, the machinery that governs this process remains poorly understood. In this study, we demonstrate that MTM1 is the main enzyme responsible for the production of phosphatidylinositol 5-phosphate, which in turn fuels PI5P 4-kinase α to produce a minor and functional pool of phosphatidylinositol 4,5-bisphosphate that concentrates in PLPs containing the scaffolding protein Tks5, Dynamin-2, and the fusogenic protein Myomaker. Collectively, our data reveal a functional crosstalk between a PI-phosphatase and a PI-kinase in the regulation of PLP formation.

Phosphoinositide 3-kinases regulate many cellular functions, including migration, growth, proliferation, and cell survival. Early studies equated the inhibition of Class I PI3Ks with loss of; phosphatidylinositol 3,4,5-trisphosphate (PIP3), but over time, it was realised that these; treatments also depleted phosphatidylinositol 3,4-bisphosphate (PI(3,4)P2). In recent years, the; use of better tools and an improved understanding of its metabolism have allowed for the; identification of specific roles of PI(3,4)P2. This includes the production of PI(3,4)P2 and the; activation of its effector Akt2 in response to growth factor signalling. In contrast, a lysosomal pool of PI(3,4)P2 is a negative regulator of mTORC1 during growth factor deprivation. A growing body of literature also demonstrates that PI(3,4)P2 controls many dynamic plasmalemmal processes. The significance of PI(3,4)P2 in cell biology is increasingly evident.

The myotubularin family, encompassing myotubularin 1 (MTM1) and 14 myotubularin-related proteins (MTMRs), represents a conserved group of phosphatases featuring a protein tyrosine phosphatase domain. Nine members are characterized by an active phosphatase domain C(X)5R, dephosphorylating the D3 position of PtdIns(3)P and PtdIns(3,5)P2. Mutations in myotubularin genes result in human myopathies, and several neuropathies including X-linked myotubular myopathy and Charcot-Marie-Tooth type 4B. MTM1, MTMR6 and MTMR14 also contribute to Ca2+ signaling and Ca2+ homeostasis that play a key role in many MTM-dependent myopathies and neuropathies. Here we explore the evolving roles of MTM1/MTMRs, unveiling their influence on critical aspects of Ca2+ signaling pathways.

The TTG2 transcription factor of Arabidopsis regulates a set of epidermal traits, including the differentiation of leaf trichomes, flavonoid pigment production in cells of the inner testa (or seed coat) layer and mucilage production in specialized cells of the outer testa layer. Despite the fact that TTG2 has been known for over twenty years as an important regulator of multiple developmental pathways, little has been discovered about the downstream mechanisms by which TTG2 co-regulates these epidermal features. In this study, we present evidence of phosphoinositide lipid signaling as a mechanism for the regulation of TTG2-dependent epidermal pathways. Overexpression of the AtPLC1 gene rescues the trichome and seed coat phenotypes of the ttg2-1 mutant plant. Moreover, in the case of seed coat color rescue, AtPLC1 overexpression restored expression of the TTG2 flavonoid pathway target genes, TT12 and TT13/AHA10. Consistent with these observations, a dominant AtPLC1 T-DNA insertion allele (plc1-1D) promotes trichome development in both wild-type and ttg2-3 plants. Also, AtPLC1 promoter:GUS analysis shows expression in trichomes and this expression appears dependent on TTG2. Taken together, the discovery of a genetic interaction between TTG2 and AtPLC1 suggests a role for phosphoinositide signaling in the regulation of trichome development, flavonoid pigment biosynthesis and the differentiation of mucilage-producing cells of the seed coat. This finding provides new avenues for future research at the intersection of the TTG2-dependent developmental pathways and the numerous molecular and cellular phenomena influenced by phospholipid signaling.

In the last three decades, the presence of phospholipids in the nucleus has been shown and thoroughly investigated. A considerable amount of interest has been raised about nuclear inositol lipids, mainly because of their role in signaling acting. Here, we review the main issues of nuclear phospholipid localization and the role of nuclear inositol lipids and their related enzymes in cellular signaling, both in physiological and pathological conditions.