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Key message SEC14L-PITPs guide membrane recognition and signaling. An increasingly complex modular structure of SEC14L-PITPs evolved in land plants compared to green algae. SEC14/CRAL-TRIO and GOLD domains govern membrane binding specificity. Abstract SEC14-like phosphatidylinositol transfer proteins (SEC14L-PITPs) provide cues for membrane identity by exchanging lipophilic substrates, ultimately governing membrane signaling. Flowering plant SEC14L-PITPs often have modular structure and are associated with cell division, development, and stress responses. Yet, structure–function relationships for biochemical–cellular interactions of SEC14L-PITPs are rather enigmatic. Here, we evaluate the phylogenetic relationships of the SEC14L-PITP superfamily in the green lineage. Compared to green algae, land plants have an extended set of SEC14L-PITPs with increasingly complex modular structure. SEC14-GOLD PITPs, present in land plants but not Chara, diverged to three functional subgroups, represented by the six PATELLIN (PATL) proteins in Arabidopsis. Based on the example of Arabidopsis PATL2, we dissect the functional domains for in vitro binding to phosphoinositides and liposomes and for plant cell membrane association. While the SEC14 domain and its CRAL-TRIO-N-terminal extension serve general membrane attachment of the protein, the C-terminal GOLD domain directs it to the plasma membrane by recognizing specific phosphoinositides. We discuss that the different domains of SEC14L-PITPs integrate developmental and environmental signals to control SEC14L-PITP-mediated membrane identity, important to initiate dynamic membrane events. Electronic supplementary material The online version of this article (10.1007/s11103-020-01067-y) contains supplementary material, which is available to authorized users.

Phospholipids regulate numerous cellular functions and their deregulation is known to be associated with cancer development. Here, we show for the first time that expression of the E6 oncoprotein of human papillomavirus type 8 (HPV8) leads to a profound increase in nuclear phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) lipid levels in monolayer cultures, that led to an aberrant phospholipidation of cellular proteins. Elevated PI(4,5)P2 levels in organotypic skin cultures, skin tumors of K14-HPV8-E6 transgenic mice as well as HPV8 positive skin carcinomas highly suggest a decisive role of PI(4,5)P2 in HPV associated squamous-cell-carcinoma development. Furthermore, mass-spectrometric analysis confirmed an increase of PI(4,5)P2, which was characterized by a shift in the distribution of lipid species. PI(4,5)P2 upregulation was independent of E6 interference with MAML1. However, E6 does interfere with the PI(4,5)P2 metabolic pathway by upregulation of phosphatidylinositol-4-phosphate-5-kinase type I and phosphatidylinositol-5-phosphate 4-kinase type II as well as the binding to 5’-phosphatase OCRL and phosphatidylinositol. All of these mechanisms combined may contribute to PI(4,5)P2 elevation in E6 positive cells. The identification of CAND1 and SND1 – two proteins known to be involved in carcinogenic processes – were significantly stronger phospholipidized in the presence of E6. In conclusion we provide evidence that the modulation of the PI(4,5)P2 metabolism is a novel oncogenic mechanism relevant for HPV-induced carcinogenesis.

Background: The transformation of the supported lipid bilayer (SLB) membrane by extracted cytosol from living resources, has recently drawn much attention. It enables us to address the question of whether the purified phospholipid SLB membrane, including lipids related to amoeba locomotion, which was discussed in many previous studies, exhibits membrane deformation in the presence of cytosol extracted from amoeba; Methods: In this report, a method for reconstituting a supported lipid bilayer (SLB) membrane, composed of purified phospholipids and cytosol extracted from Dictyostelium discoideum, is described. This technique is a new reconstitution method combining the artificial constitution of membranes with the reconstitution using animate cytosol (without precise purification at a molecular level), contributing to membrane deformation analysis; Results: The morphology transition of a SLB membrane composed of phosphatidylcholines, after the addition of cytosolic extract, was traced using a confocal laser scanning fluorescence microscope. As a result, pore formation in the SLB membrane was observed and phosphatidylinositides incorporated into the SLB membrane tended to suppress pore formation and expansion; Conclusions: The current findings imply that phosphatidylinositides have the potential to control cytoplasm activity and bind to a phosphoinositide-containing SLB membrane.

The regulation of animal longevity shows remarkable plasticity, in that a variety of genetic lesions are able to extend lifespan by as much as tenfold. Such studies have implicated several key signaling pathways that must normally limit longevity, since their disruption prolongs life. Little is known, however, about the proximal effectors of aging on which these pathways are presumed to converge, and to date, no pharmacologic agents even approach the life-extending effects of genetic mutation. In the present study, we have sought to define the downstream consequences of age-1 nonsense mutations, which confer 10-fold life extension to the nematode C. elegans ― the largest effect documented for any single mutation. Such mutations insert a premature stop codon upstream of the catalytic domain of the AGE-1/ p110α subunit of class-I PI3K. As expected, we do not detect class-I PI3K (and based on our sensitivity, it constitutes PIA6>PIA12) as lifespan. A second reporter, PEPCK::GFP, was equally activated (~40%) by all three.

Profilin is an ubiquitous G-actin binding protein in eukaryotic cells. Lack of both profilin isoforms in Dictyostelium discoideum resulted in impaired cytokinesis and an arrest in development. A restriction enzyme–mediated integration approach was applied to profilin-minus cells to identify suppressor mutants for the developmental phenotype. A mutant with wild-type–like development and restored cytokinesis was isolated. The gene affected was found to code for an integral membrane glycoprotein of a predicted size of 88 kD containing two transmembrane domains, one at the NH2 terminus and the other at the COOH terminus. It is homologous to mammalian CD36/LIMP-II and represents the first member of this family in D. discoideum, therefore the name DdLIMP is proposed. Targeted disruption of the lmpA gene in the profilin-minus background also rescued the mutant phenotype. Immunofluorescence revealed a localization in vesicles and ringlike structures on the cell surface. Partially purified DdLIMP bound specifically to PIP2 in sedimentation and gel filtration assays. A direct interaction between DdLIMP and profilin could not be detected, and it is unclear how far upstream in a regulatory cascade DdLIMP might be positioned. However, the PIP2 binding of DdLIMP points towards a function via the phosphatidylinositol pathway, a major regulator of profilin.

Background The cytosol of amoeba cells controls the membrane deformation during their motion in vivo. To investigate such ability of the cytosol of amoeba cell, Dictyostelium discoideum (Dictyostelium), in vitro, we used lipids extracted from Dictyostelium and commercially available phospholipids, and prepared substrate-supported lipid membrane patches on the micrometer scale by spin coating. Results We found that the spin coater holder, which has pores (pore size = 3.1 mm) of negative pressure to hold the cover glass induced the concave surface of the cover glass. The membrane lipid patches were formed at each position in the vicinity of the holder pores and their sizes were in the range of 2.7 to 3.2 × 104 μm2. After addition of the cytosol extracted from Dictyostelium to the lipid membrane patches, through time-lapse observation with a confocal laser scanning fluorescence microscope, we observed an autonomous buckling of the Dictyostelium lipid patches and localized behaviours of proteins found within. Conclusion The current method serves as the novel technique for the preparation of film patches in which the positions of patches are controlled by the holder pores without fabricating, modifying, and arranging the chemical properties of the solution components of lipids. The findings imply that lipid-binding proteins in the cytosol were adsorbed and accumulated within the Dictyostelium lipid patches, inducing the transformation of the cell-sized patch. Electronic supplementary material The online version of this article (doi:10.1186/1754-1611-9-3) contains supplementary material, which is available to authorized users.