Biology subjects

Alessandrini, A.

Publications and source records attributed to Alessandrini, A..

3 recordsLinked to original sources

bioRxiv2026

Metabolic reprogramming controlled by NF-YA alternative splicing creates therapeutic opportunities in colorectal cancer

Metabolic reprogramming is a fundamental strategy that allows colorectal cancer (CRC) cells to endure microenvironmental constraints and sustain malignant progression. Here, we identify the transcription factor NF-Y as a master regulator of glutamine metabolism in CRC, with particular relevance to the aggressive CMS4 subtype. Loss of function experiments, integrated with metabolomic and transcriptomic analyses, reveal a critical role for NF-YA in regulating glutamine metabolism in CRC cells. Complementary gain of function studies pinpoint NF-YAl as the isoform specifically driving glutamine-centered rewiring. Mechanistically, NF-YAl directly binds the Glul promoter, inducing transcriptional upregulation of glutamine synthetase and increasing intracellular glutamine availability. This metabolic reprogramming enhances resistance to mechanical shear and oxidative stress under glutamine-limiting conditions, thereby promoting migratory and metastatic traits. Importantly, pharmacological inhibition of glutamine synthesis, but not uptake or downstream catabolism, selectively abrogates the survival and migratory advantage of NF-YAlhigh cells both in vitro and in vivo, highlighting a targetable vulnerability in aggressive CRC. Beyond CRC cell-autonomous advantage, NF-YAl-dependent glutamine biosynthesis reshapes the tumor microenvironment by promoting M2 macrophage polarization. Conditioned medium from NF-YAlhigh CRC cells is sufficient to induce human monocytes to adopt an M2-like phenotype. This effect is dependent on NF-YAlhigh tumor-derived glutamine, as inhibition of glutamine uptake by monocytes fully blocks their conversion to M2. In line with this, integrative analyses of patient-derived datasets underscore the predictive relevance of the NF-YAl-Glul-M2 axis in driving CRC aggressiveness. These findings define glutamine synthetase as a pivotal mediator of NF-YAl activity and a promising druggable metabolic Achilles heel in NF-YAlhigh CRC tumors.

cancer biology↗

bioRxiv2026

Endothelial PTBP1 Deletion in Transplanted Cardiac Tissue Limits Cardiac Allograft Vasculopathy

BackgroundCardiac allograft vasculopathy (CAV) is a leading cause of late graft failure and mortality following heart transplantation, with limited therapeutic options. Endothelial cells (ECs), at the interface between the donor graft and host immune system, play a central role in CAV development. However, the molecular mechanisms driving endothelial dysfunction and vascular remodeling in chronic heart transplant rejection remain poorly understood. MethodsTo characterize endothelial alterations associated with CAV, we isolated nuclei from cardiac tissues of four human donor groups: (1) early post-transplant CAV-negative surveillance biopsies, (2) CAV-negative explanted grafts with acute cellular rejection (ACR), (3) late-stage CAV-positive explanted grafts, and (4) naive non-transplanted control hearts. We applied intranuclear cellular indexing of transcriptomes and epitopes (inCITE-seq) to profile endothelial gene expression together with nuclear protein levels of splice factor polypyrimidine tract-binding protein 1 (PTBP1), a key post-transcriptional regulator of endothelial inflammatory responses. Functional relevance of PTBP1 was assessed using endothelial-specific deletion of Ptbp1 in an F1 hybrid murine model of CAV. ResultsIn human CAV, endothelial cells exhibited increased transforming growth factor-{beta} (TGF-{beta}) signaling and reduced oxidative phosphorylation (OxPhos) transcripts. Nuclear PTBP1 protein levels were markedly elevated in CAV endothelium and were associated with TGF-{beta}-responsive transcriptional programs and correlated with clinical indices of cardiac dysfunction. In murine heart transplants, endothelial-specific deletion of Ptbp1 markedly reduced hallmarks of CAV, including neointimal hyperplasia, fibrosis, and lymphocyte activation. At the molecular level, endothelial Ptbp1 deletion prevented suppression of mitochondrial transcripts and preserved mitochondrial content and integrity under hypoxic stress, attenuating interferon signaling in endothelial cells. ConclusionThese findings identify PTBP1 as a central endothelial regulator linking pro-fibrotic stress to mitochondrial dysfunction and immune activation in chronic cardiac allograft rejection. Targeting endothelial PTBP1 may represent a strategy to limit chronic graft injury while minimizing systemic immunosuppression.

immunology↗

bioRxiv2024

The CDKL5 kinase undergoes liquid-liquid phase separation driven by a serine-rich C-terminal region and impaired by neurodevelopmental disease-related truncations

Mutations of the cyclin-dependent kinase-like 5 (CDKL5) gene, which encodes a serine/threonine protein kinase, can cause the CDKL5 deficiency disorder (CDD), a severe neurodevelopmental disease characterized by epileptic encephalopathy and neurocognitive impairment. The CDKL5 kinase consists of a catalytic N-terminal domain (NTD) and a less characterized C-terminal domain (CTD). Numerous disease-related mutations truncate CDKL5, leaving the NTD intact while variably shortening the CTD, which highlights the importance of the CTD for CDKL5 function. By systematically analyzing CDKL5 compositional features and evolutionary dynamics, we found that the CTD is a low-complexity region (LCR) highly enriched in serine residues and with a high propensity to undergo liquid-liquid phase separation (LLPS), a biophysical process of condensation controlling protein localization and function. Using a combination of super-resolution imaging, electron microscopy, and molecular and cellular approaches, including optogenetic LLPS induction, we discovered that CDKL5 undergoes LLPS, predominantly driven by its CTD, forming membraneless condensates in neuronal and non-neuronal cells. A CTD internal fragment (CTIF) plays a pivotal LLPS-promoting role, along with the distal portion of the protein. Indeed, two disease-related truncating mutations (S726X and R781X), eliding variable portions of the CTIF, significantly impair LLPS. This impairment is paralleled at the functional level by a reduction in the CDKL5-dependent phosphorylation of EB2, a known CDKL5 target. These findings demonstrate that CDKL5 undergoes LLPS, driven by a CTD region elided by most disease-related truncating mutations. Its loss--through the impairment of CDKL5 LLPS and functional activity--may play a key role in the molecular pathogenesis of CDD.

molecular biology↗