bims-toxgon 2026-08-23 papers

Issue of 2026–08–23
five papers selected by
Lakesh Kumar, BITS Pilani

bioRxiv. 2026 Aug 05. pii: 2026.08.05.743015. [Epub ahead of print]

A proximity biotinylation approach for the identification of membrane contact site proteins in Toxoplasma gondii.

Inter-organellar communication is crucial for cellular function. Inside the cell, organelles interact with each other via membrane contact sites (MCSs). These structures mediate the close apposition of two organellar membranes to allow for the exchange of metabolites, ions and lipids. Most of what is known about MCSs comes only from a handful of well-studied metazoans, particularly yeast and mammals. Apicomplexans are parasites that drive human disease throughout the world. Yet, little is known about the makeup or function of their MCSs, leaving a gap in our understanding of how organelles communicate beyond conventional model eukaryotes. Here, we used a proximity biotinylation approach to map the surface proteome of three organelles in the model apicomplexan Toxoplasma gondii : the apicoplast-a non-photosynthetic plastid found only in apicomplexans-its single mitochondrion and the endoplasmic reticulum. By subtracting a cytosolic spatial reference, our high-stringency proteomic analysis uncovered candidate proteins localized simultaneously to multiple organellar surfaces suggesting their role as MCS components. We then validate our approach by characterizing a candidate involved in the association between the apicoplast and the mitochondrion. Overall, our findings provide a valuable approach to identify MCSs in apicomplexans and set the stage to apply our approach to other organelles in these pathogens.
Highlights: Generation of surface proteomes for the apicoplast, mitochondrion, and ER in Toxoplasma gondii Mapping of the first endoplasmic reticulum and mitochondrial surface proteomes in T. gondii Identified novel membrane contact site candidate proteinsValidated a membrane contact site candidate mediating mitochondrion-apicoplast interactions.

DOI: https://doi.org/10.64898/2026.08.05.743015

PLoS Pathog. 2026 Aug;22(8): e1014526

The malaria parasite Plasmodium falciparum Vps15 is a Phosphatidylinositol 3-kinase interactor important for host-cell cytosol delivery to the food vacuole and apicoplast and mitochondrion dynamics.

Alexandra Bourgeois, Louis-David Martin, Harpreet Kaur, Dominic Gagnon, Joel B Dacks, Dave Richard.

The malaria parasite Plasmodium falciparum is an obligate intracellular organism that spends an important part of its lifecycle inside human erythrocytes. The endocytosis of host-cell cytosol and its delivery to a lysosome-like organelle called the food vacuole are critical for the parasite's survival and proliferation. Recent work has started to identify some of the molecular players involved in this pathway, but much remains to be discovered. Evidence suggests that phosphatidylinositol-3-phosphate (PI3P) plays a central role in this process. In unicellular eukaryotes, such as yeast, PI3P is generated by a single PI3-kinase, whose activity is regulated by a pseudokinase called Vps15. P. falciparum also possesses a PI3K that generates PI3P and bioinformatics analysis has revealed the presence of an uncharacterized putative orthologue of Vps15. We here present our characterization of PfVps15. We first show that it is constitutively expressed throughout the asexual erythrocytic cycle and that it interacts with PfPI3K, but unlike in yeast and mammalian cells, it is potentially not part of a heterotetrameric complex. The removal of PfVps15 from its site of action by knock sideways led to rapid parasite death. Phenotypic analyses revealed a decrease in PI3P levels, the abrogation of the delivery of host-cell cytosol containing vesicles to the food vacuole, and defects in apicoplast biogenesis and mitochondrial fission. Collectively, our data has identified a protein critical for the synthesis of PI3P and provides molecular evidence for the importance of this lipid in the vesicular trafficking pathway of host-cell cytosol, and apicoplast and mitochondrion dynamics.

DOI: https://doi.org/10.1371/journal.ppat.1014526

Methods Enzymol. 2026 ;pii: S0076-6879(26)00159-X. [Epub ahead of print]733 1-16

Experimental strategies for evaluating the impact of HDACs and HDAC inhibitors on the epigenome. ChiP-seq.

Martina Minisini, Alessio Bertozzo, Eros Di Giorgio, Claudio Brancolini.

HDACs (histone deacetylases) are components of multiprotein complexes that remove the acetyl group from lysine residues (and other acyl groups) to regulate protein function. This important post-translational modification can affect the local hydrophobic environment of a specific protein surface. When the protein substrates are histones, the regulation of lysine acetylation influences chromatin accessibility and gene expression. Defining the different genomic regions under the influence of specific HDACs is fundamental to understanding how these enzymes control the epigenetic status of cells. Furthermore, it is important to clarify the mechanisms of action of HDAC inhibitors from a therapeutic perspective. ChIP-seq (Chromatin Immunoprecipitation followed by sequencing) is a powerful technique to identify where histone acetylations regulated by HDACs or influenced by HDAC inhibitors occur in the genome. Here, we describe a protocol to perform a ChIP-seq experiment to detect the genome-wide distribution of histone acetylation.

Keywords: ChIP-seq; H3K27ac; HDACi; HDACs; Normalization; Sonication

DOI: https://doi.org/10.1016/bs.mie.2026.05.031

Methods Enzymol. 2026 ;pii: S0076-6879(26)00166-7. [Epub ahead of print]733 17-31

Methods for the kinetic analysis of time-dependent HDAC6 inhibitors.

Edoardo Cellupica, Gianluca Fossati, Christian Steinkühler, Gianluca Caprini.

The class IIb histone deacetylase HDAC6 is responsible for the deacetylation of multiple cytoplasmic substrates, including α-tubulin and Miro-1, and has emerged as an important therapeutic target in neuropathies. In this context, several HDAC6 inhibitors effective in neuropathic models, exhibit slow-binding and slow-release behavior, thereby requiring kinetic approaches that extend beyond commonly used endpoint assays, which can lead to misinterpretation of inhibitor potency and mechanism of action. Notably, compounds with slow dissociation kinetics may offer improved therapeutic properties in neuropathic conditions, as sustained target engagement can translate into prolonged improvement of axonal transport and cytoskeletal dynamics. In this chapter, we outline a comprehensive methodology for the study of HDAC6 inhibition and the identification of slow-binding and slow-dissociating inhibitors.

Keywords: Difluoromethyl-1,3,4-oxadiazoles; HDAC6; Mechanistic enzymology; Neuropathy; Residence time; Slow-binding inhibition

DOI: https://doi.org/10.1016/bs.mie.2026.05.038

FEBS Open Bio. 2026 Aug 19.

Threonine 348 regulates the subcellular localization of PTEN.

Takashi Kato, Suzu Tanaka, Miyu Ohashi.

The tumor suppressor PTEN (phosphatase and tensin homolog) dephosphorylates PIP3 (phosphatidylinositol (3,4,5)-trisphosphate) at the plasma membrane and protects genomic integrity in the nucleus; thus, regulation of PTEN subcellular localization is crucial. Previous studies have shown that the PTEN350 fragment is markedly enriched in the nucleus, a feature not explained by the N-terminal nuclear localization signal. In this study, we generated PTEN fragments of various lengths and identified PTEN348 (residues 1-348), which showed prominent nuclear localization. Furthermore, the replacement of threonine 348 (Thr348) with other amino acids reduced the nuclear localization of the PTEN348 and PTEN350 fragments. Moreover, we found that PTENA4 and PTENK13R,A4 localized predominantly to the nucleus and plasma membrane, respectively, and that substitution of Thr348 with aspartic acid resulted in cytoplasmic localization in both mutants. Collectively, these results indicate that Thr348 is a key contributor to the regulation of PTEN subcellular localization.

Keywords: C2 domain; PTEN; nucleus; plasma membrane; subcellular localization; threonine 348

DOI: https://doi.org/10.1002/2211-5463.70328