Biomedical subjects

Galina Tuymetova

Publications and source records attributed to Galina Tuymetova.

4 recordsLinked to original sources

PubMed2023

SARS-CoV-2 ORF3a expression in brain disrupts the autophagy-lysosomal pathway, impairs sphingolipid homeostasis, and drives neuropathogenesis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causes injury to multiple organ systems, including the brain. SARS-CoV-2's neuropathological mechanisms may include systemic inflammation and hypoxia, as well as direct cell damage resulting from viral infections of neurons and glia. How the virus directly causes injury to brain cells, acutely and over the long term, is not well understood. In order to gain insight into this process, we studied the neuropathological effects of open reading frame 3a (ORF3a), a SARS-CoV-2 accessory protein that is a key pathological factor of the virus. Forced ORF3a brain expression in mice caused the rapid onset of neurological impairment, neurodegeneration, and neuroinflammation-key neuropathological features found in coronavirus disease (COVID-19, which is caused by SARS-CoV-2 infection). Furthermore, ORF3a expression blocked autophagy progression in the brain and caused the neuronal accumulation of α-synuclein and glycosphingolipids, all of which are linked to neurodegenerative disease. Studies with ORF3-expressing HeLa cells confirmed that ORF3a disrupted the autophagy-lysosomal pathway and blocked glycosphingolipid degradation, resulting in their accumulation. These findings indicate that, in the event of neuroinvasion by SARS-CoV-2, ORF3a expression in brain cells may drive neuropathogenesis and be an important mediator of both short- and long-term neurological manifestations of COVID-19.

Animals↗

PubMed2005

A plasma membrane pool of phosphatidylinositol 4-phosphate is generated by phosphatidylinositol 4-kinase type-III alpha: studies with the PH domains of the oxysterol binding protein and FAPP1.

The PH domains of OSBP and FAPP1 fused to GFP were used to monitor PI(4)P distribution in COS-7 cells during manipulations of PI 4-kinase (PI4K) activities. Both domains were associated with the Golgi and small cytoplasmic vesicles, and a small fraction of OSBP-PH was found at the plasma membrane (PM). Inhibition of type-III PI4Ks with 10 microM wortmannin (Wm) significantly reduced but did not abolish Golgi localization of either PH domains. Downregulation of PI4KIIalpha or PI4KIIIbeta by siRNA reduced the localization of the PH domains to the Golgi and in the former case any remaining Golgi localization was eliminated by Wm treatment. PLC activation by Ca2+ ionophores dissociated the domains from all membranes, but after Ca2+ chelation, they rapidly reassociated with the Golgi, the intracellular vesicles and with the PM. PM association of the domains was significantly higher after the Ca2+ transient and was abolished by Wm pretreatment. PM relocalization was not affected by down-regulation of PI4KIIIbeta or -IIalpha, but was inhibited by down-regulation of PI4KIIIalpha, or by 10 microM PAO, which also inhibits PI4KIIIalpha. Our data suggest that these PH domains detect PI(4)P formation in extra-Golgi compartments under dynamic conditions and that various PI4Ks regulate PI(4)P synthesis in distinct cellular compartments.

1-Phosphatidylinositol 4-Kinase↗

PubMed2002

Inositol lipid binding and membrane localization of isolated pleckstrin homology (PH) domains. Studies on the PH domains of phospholipase C delta 1 and p130.

The relationship between the ability of isolated pleckstrin homology (PH) domains to bind inositol lipids or soluble inositol phosphates in vitro and to localize to cellular membranes in live cells was examined by comparing the PH domains of phospholipase Cdelta(1) (PLCdelta(1)) and the recently cloned PLC-like protein p130 fused to the green fluorescent protein (GFP). The prominent membrane localization of PLCdelta(1)PH-GFP was paralleled with high affinity binding to inositol 1,4,5-trisphosphate (InsP(3)) as well as to phosphatidylinositol 4,5-bisphosphate-containing lipid vesicles or nitrocellulose membrane strips. In contrast, no membrane localization was observed with p130PH-GFP despite its InsP(3) and phosphatidylinositol 4,5-bisphosphate-binding properties being comparable with those of PLCdelta(1)PH-GFP. The N-terminal ligand binding domain of the type I InsP(3) receptor also failed to localize to the plasma membrane despite its 5-fold higher affinity to InsP(3) than the PH domains. By using a chimeric approach and cassette mutagenesis, the C-terminal alpha-helix and the short loop between the beta6-beta7 sheets of the PLCdelta(1)PH domain, in addition to its InsP(3)-binding region, were identified as critical components for membrane localization in intact cells. These data indicate that binding to the inositol phosphate head group is necessary but may not be sufficient for membrane localization of the PLCdelta(1)PH-GFP fusion protein, and motifs located within the C-terminal half of the PH domain provide auxiliary contacts with additional membrane components.

Amino Acid Sequence↗

PubMed2002

Characterization of type II phosphatidylinositol 4-kinase isoforms reveals association of the enzymes with endosomal vesicular compartments.

Phosphorylation of phosphatidylinositol (PI) to PI 4-phosphate is one of the key reactions in the production of phosphoinositides, lipid regulators of several cellular functions. This reaction is catalyzed by multiple enzymes that belong either to the type II or the type III family of PI 4-kinases. Type III enzymes are structurally similar to PI 3-kinases and are sensitive to PI 3-kinase inhibitors. In contrast, the recent cloning of the first type II PI 4-kinase enzyme defined a novel enzyme family. Here we characterize a new member of this family, the type IIbeta enzyme that has been identified in the NCBI data base based on its homology to the first-cloned type IIalpha enzyme. The type IIbeta enzyme has a primary transcript size of approximately 3.8 kb and shows wide tissue distribution. It contains an open reading frame of 1.4 kb, encoding a protein of approximately 54 kDa. Sequence comparison reveals a high degree of similarity to the type IIalpha enzyme within the C-terminal catalytic domain but significantly lower homology within the N-terminal region. Expression of both enzyme yields increased PI 4-kinase activity that is associated with the microsomal membrane fractions and is significantly lower for the type IIbeta than the type IIalpha form. Both enzymes use PI as their primary substrate and have no detectable activity on PI monophosphates. Epitope-tagged as well as green fluorescent protein-tagged forms of both enzymes localize primarily to intracellular membranes and show prominent co-localization with early endosomes and recycling endosomes but not with the Golgi. These compartments participate in the processing of both the transferrin receptor and the G protein-coupled AT(1A) angiotensin receptor. Our data indicate the existence of multiple forms of type II PI 4-kinase in mammalian cells and suggest that their functions are related to the endocytic pathway.

1-Phosphatidylinositol 4-Kinase↗