Biology subjects

Abdul-Salam, V. B.

Publications and source records attributed to Abdul-Salam, V. B..

6 recordsLinked to original sources

bioRxiv2025

Mechanistic Elucidation of CLIC1 Membrane Insertion via Structural and Dynamic Modulation

Chloride Intracellular Channel 1 (CLIC1) is a metamorphic protein capable of transitioning from a soluble cytoplasmic state to a membrane-bound chloride channel. This conformational shift, crucial for physiological processes such as cell volume regulation, electrical excitability, and angiogenesis, is linked to pathological conditions including malignancies and cardiovascular diseases. Despite its significance, the molecular mechanism driving CLIC1s membrane insertion has remained elusive. Using an integrated structural biology approach combining NMR spectroscopy, SAXS, biophysical methods, and mutagenesis, we uncover the dynamic landscape underpinning CLIC1 function. Solution NMR and SAXS reveal that CLIC1 adopts a conformational ensemble in equilibrium, characterized by a compact ground state and a partially extended state exposing key membrane-interacting regions. Zn2+ binding acts as a critical trigger, inducing structural rearrangements, increasing protein flexibility, and promoting oligomerization essential for membrane insertion. Our findings demonstrate that structural flexibility, particularly within dynamic loop regions and interdomain linkers, is intrinsic to CLIC1s ability to adapt to membrane interactions. Zn2+-induced dimerization and tetramerization were identified as key steps preceding insertion, with mutations in the transmembrane (TM) region revealing pivotal roles for residues R29 and W35 in modulating protein dynamics, oligomerization and insertion eXiciency. This study provides a mechanistic framework for CLIC1s transition to its membrane-bound state, oXering insights into the interplay between conformational dynamics, oligomerization, and metal ion modulation. These findings pave the way for targeted strategies to regulate CLIC1 activity in pathological conditions, underscoring its potential as a therapeutic target.

biochemistry↗

bioRxiv2022

Structure-based discovery and in vitro validation of selective inhibitors of Chloride Intracellular Channel 4 protein.

Chloride Intracellular Channel Protein 4 (CLIC4) is a novel class of intracellular ion channel highly implicated in tumour and vascular biology. It regulates cell proliferation, apoptosis and angiogenesis; and is involved in multiple pathologic signaling pathways. Absence of specific inhibitors impedes its advancement to translational research. Here, we integrate structural bioinformatics and experimental research approach for the discovery and validation of small-molecule inhibitors of CLIC4. High-affinity allosteric binders were identified from a library of 1615 Food and Drug Administration (FDA)-approved drugs via a high-performance computing-powered blind-docking approach, resulting in the selection of amphotericin B and rapamycin. NMR assays confirmed the binding of the drugs. Both drugs reversed stress-induced membrane translocation of CLIC4 and inhibited endothelial cell migration. Structural and dynamics simulation studies further revealed that the inhibitory mechanisms of these compounds were hinged on the allosteric modulation of the catalytic glutathione (GSH)-like site loop and the extended catalytic {beta} loop which may elicit interference with the catalytic activities of CLIC4. Structure-based insights from this study provide the basis for the selective targeting of CLIC4 to treat the associated pathologies.

molecular biology↗

bioRxiv2021

Zn2+ triggered two-step mechanism of CLIC1 membrane insertion and activation into chloride channels

The CLIC protein family displays the unique feature of altering its structure from a soluble form to a membrane-bound chloride channel. CLIC1, a member of this family, is found in the cytoplasm or in internal and the plasma membranes, with membrane relocalisation linked to endothelial disfunction, tumour proliferation and metastasis. The molecular switch promoting CLIC1 activation remains unclear. Here, cellular chloride efflux assays and immunofluorescence microscopy studies have identified Zn2+ intracellular release as the trigger for CLIC1 activation and membrane relocalisation. Biophysical assays confirmed specific binding to Zn2+, inducing membrane association and enhancing chloride efflux in a pH dependent manner. Together, our results identify a two-step mechanism with Zn2+ binding as the molecular switch promoting CLIC1 membrane insertion, followed by pH activation of chloride efflux.

biochemistry↗

bioRxiv2020

NOVEL NUCLEAR ROLE OF HDAC6 IN PROGNOSIS AND THERAPEUTIC TARGET FOR COLORECTAL CANCER

Histone deacetylase 6 (HDAC6) inhibition is a potential treatment of a wide range of cancer types via the acetylation of diverse proteins in the cytoplasm. However, the regulation of histone acetylation and the maintenance of higher-order chromatin structure remains unidentified. Here, we investigated the effect of selective inhibition of HDAC6 by histone acetylation, chromatin relaxation assays, co-immunoprecipitation, acetylome peptide array and in vivo RNA microarray. Our data shows that nuclear HDAC6 physically interacts with the Histone 4 lysine 12 residue, and that HDAC6 inhibition increases acetylation specifically at this residue in several cancer types. Inhibition induces major chromatin structure modulation, but has no equivalent effect on knockout HDAC6-/- MEF cells. We identified several novel HDAC6-deacetylated substrates and high expression of HDAC6 in colorectal cancer (CRC) tissue association with reduced levels of H4K12ac and independent of the key CRC driver mutations, but positively associated with EGFR expression. Furthermore, in vivo HDAC6 inhibition induces significant tumor regression in a CRC xenograft mice model with significant changes in the expression of functional nuclear genes. We also demonstrated that a DNA damaging agent in combination with selective HDAC6 inhibition is effective and acts synergistically, inducing chromatin relaxation and increased cell death in CRC cells. CRC tissues (Normal versus tumor; n=58 matched pairs) together with TCGA data analysis of 467 CRC patients showed that high HDAC6 expression is associated with metastasis, overall and disease-free survival, and is an independent risk factor of CRC stage progression. Our findings designate a new role for nuclear HDAC6 both in cancer prognosis and as a new therapeutic target for CRC and other types of cancer. HighlightHistone deacetylases 6 activity; Chromatin relaxation; Histone modifications; Gene array; DOX: doxorubicin; OXA: oxaliplatin; 5-FU: fluorouracil; Ac: acetylation; MNase: Micrococal nuclease.

cancer biology↗

bioRxiv2020

Role of endothelial microRNA-150 in pulmonary arterial hypertension

Endothelial dysfunction contributes to the vascular pathology in pulmonary arterial hypertension (PAH). Circulating levels of endothelial miR-150 are reduced in PAH and act as an independent predictor of patient survival. The role of endothelial miR-150 in vascular dysfunction in PAH is not well understood. Endothelium-targeted miR-150 delivery prevented the disease in Sugen/hypoxia mice, while endothelial knockdown of miR-150 had adverse effects. miR-150 target genes revealed significant associations with PAH pathways, including proliferation, inflammation and phospholipid signaling, with PTEN-like mitochondrial phosphatase (PTPMT1) most markedly altered. PTPMT1 reduced inflammation, apoptosis and improved mitochondrial function in human pulmonary endothelial cells and blood-derived endothelial colony forming cells (ECFCs) from idiopathic PAH. Beneficial effects of miR-150 in vitro and in vivo were linked with PTPMT1-dependent biosynthesis of mitochondrial phospholipid cardiolipin and reduced expression of pro-apoptotic, pro-inflammatory and pro-fibrotic genes, including c-MYB, NOTCH3, TGF-{beta} and Col1a1. In conclusion, we are first to show that miR-150-PTPMT1-cardiolipin pathway attenuates pulmonary endothelial damage induced by vascular stresses and may be considered as a potential therapeutic strategy in PAH.

physiology↗

bioRxiv2019

Krüppel-like factor 2- induced microRNAs: implications for treatment of pulmonary hypertension

Flow-activated transcription factor Kruppel-like factor 2 (KLF2) signaling is compromised in pulmonary arterial hypertension (PAH). We aimed to identify KLF2-induced endothelium-protective exosomal microRNAs of potential therapeutic significance.\n\nEight exosomal microRNAs elevated by KLF2 but reduced in PAH were transfected into human pulmonary artery endothelial cells. Of these, only miR-181a-5p and miR-324-5p had anti-apoptotic, anti-inflammatory and anti-proliferative effect on endothelial cells and reduced proliferation of vascular smooth muscle cells in vitro. RNA sequencing of miRNA-transfected HPAECs revealed reduced expression of multiple genes implicated in vascular remodelling, including ETS-1, NOTCH4, ACTA2, TNF-, IL-1, MMP10, MAPK and NFATC2. KLF2, miR-181a-5p and miR-324-5p were reduced, while their target genes were elevated in blood-derived endothelial colony forming cells and lung tissues from idiopathic and heritable PAH patients with disabling KLF2 mutation and Sugen/hypoxia mice. Supplementation of miR-181a-5p and miR-324-5p or silencing of their target genes attenuated proliferative and angiogenic responses in endothelial cells from idiopathic PAH and prevented development of pulmonary hypertension in Sugen/hypoxia mice.\n\nThis study highlights potential therapeutic role of KLF2-induced exosomal microRNAs in PAH.

cell biology↗