Biology subjects

Askenazi, M.

Publications and source records attributed to Askenazi, M..

6 recordsLinked to original sources

bioRxiv2026

Integrated Interactomics Reveals Novel Protein Associations: The FOXA1-PBX1 Complex as a Case Study

Protein-protein interactions (PPIs) are fundamental to cellular signaling networks, yet many remain undetected due to technical limitations of individual affinity purification approaches. To address this, we systematically mapped the interaction landscapes of six regulatory proteins involved in cell proliferation, immunity, and inflammation, including three transcription factors (TFs) and three kinases. We implemented an integrated proteomics workflow that combined four complementary affinity purification strategies: native immunoprecipitation, two crosslinking-assisted capture methods, and proximity labeling. Combining these approaches revealed distinct yet overlapping interaction profiles, uncovered numerous previously unreported interactors not reliably detected by individual methods, and robustly recovered known interactions while substantially extending PPI networks. Despite method-specific differences at the protein level, functional enrichment analyses showed strong convergence on coherent biological pathways. Biochemical approaches validated most of the previously unreported interactions, including putative weak and transient complexes stabilized by crosslinking. Functional assays revealed a previously unrecognized physical interaction between FOXA1 and PBX1 TFs and demonstrated their cooperative regulation of transcriptional programs and cell fitness in estrogen receptor (ER) positive breast cells. We propose that the FOXA1-PBX1 complex could represent a higher-order regulatory node integrating chromatin accessibility and ER-driven transcriptional output. HIGHLIGHTSO_LIComplementary affinity purification strategies uncover putative weak and transient protein-protein interactions C_LIO_LIFunctional pathway convergence validates biologically coherent interactome expansion C_LIO_LIBiochemical validations confirm unreported interactions C_LIO_LIFunctional validation studies identify a FOXA1-PBX1 pioneer factor complex that regulates estrogen receptor transcriptional programs C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=191 HEIGHT=200 SRC="FIGDIR/small/738938v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@193cc1borg.highwire.dtl.DTLVardef@3d068dorg.highwire.dtl.DTLVardef@791d5corg.highwire.dtl.DTLVardef@1769ce7_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2024

D-type cyclins regulate DNA mismatch repair in the G1 and S phases of the cell cycle, maintaining genome stability

The large majority of oxidative DNA lesions occurring in the G1 phase of the cell cycle are repaired by base excision repair (BER) rather than mismatch repair (MMR) to avoid long resections that can lead to genomic instability and cell death. However, the molecular mechanisms dictating pathway choice between MMR and BER have remained unknown. Here, we show that, during G1, D-type cyclins are recruited to sites of oxidative DNA damage in a PCNA- and p21-dependent manner. D-type cyclins shield p21 from its two ubiquitin ligases CRL1SKP2 and CRL4CDT2 in a CDK4/6-independent manner. In turn, p21 competes through its PCNA-interacting protein degron with MMR components for their binding to PCNA. This inhibits MMR while not affecting BER. At the G1/S transition, the CRL4AMBRA1-dependent degradation of D-type cyclins renders p21 susceptible to proteolysis. These timely degradation events allow the proper binding of MMR proteins to PCNA, enabling the repair of DNA replication errors. Persistent expression of cyclin D1 during S-phase increases the mutational burden and promotes microsatellite instability. Thus, the expression of D-type cyclins inhibits MMR in G1, whereas their degradation is necessary for proper MMR function in S. One-Sentence SummaryTo maintain genome stability, D-type cyclins limit mismatch repair (MMR) in G1, whereas their degradation is necessary for proper MMR function in S phase.

cell biology↗

bioRxiv2023

Compilation of all known protein changes in the human Alzheimer's disease brain

Proteomic studies of human Alzheimers disease brain tissue have exceptional potential to identify protein changes that drive disease and to identify new drug targets. Here, we detail a combined analysis of 38 published Alzheimers disease proteomic studies, generating a comprehensive map of protein changes in human brain tissue across thirteen brain regions, three disease stages (preclinical Alzheimers disease, mild cognitive impairment, advanced Alzheimers disease), and proteins enriched in amyloid plaques, neurofibrillary tangles, and cerebral amyloid angiopathy. Our dataset is compiled into a user-friendly, searchable database called NeuroPro. Our combined analysis included 18,119 reported protein differences in human Alzheimers disease brain tissue, which mapped to 5,311 total altered proteins. Proteomic studies were remarkably consistent. 848 proteins were consistently altered in [≥]5 studies, many of which are understudied in the Alzheimers field. Comparison of protein changes in early-stage and advanced Alzheimers disease revealed significant synapse, vesicle, and lysosomal changes early in disease, but widespread mitochondrial changes only in advanced Alzheimers disease. Comparison of vulnerable and resistant brain regions suggested that protein changes in resistant regions in advanced Alzheimers disease are similar to those in vulnerable regions in early-stage Alzheimers disease, indicating a temporal progression of protein dysfunction during Alzheimers disease advancement. We conclude that NeuroPro is a powerful new resource that provides new insights into human Alzheimers disease brain protein changes and highlights novel proteins of particular interest that may mechanistically drive Alzheimers disease.

neuroscience↗

bioRxiv2023

Cell wall proteomics in live Mycobacterium tuberculosis uncovers exposure of ESX substrates to the periplasm

The cell wall of mycobacteria plays a key role in interactions with the environment and its ability to act as a selective filter is crucial to bacterial survival. Proteins in the cell wall enable this function by mediating the import and export of diverse metabolites from ions to lipids to proteins. Accurately identifying cell wall proteins is an important step in assigning function, especially as many mycobacterial proteins lack functionally characterized homologues. Current methods for protein localization have inherent limitations that reduce accuracy. Here we showed that protein tagging by the engineered peroxidase APEX2 within live Mycobacterium tuberculosis enabled the accurate identification of the cytosolic and cell wall proteomes. Our data indicate that substrates of the virulence-associated Type VII ESX secretion system are exposed to the Mtb periplasm, providing insight into the currently unknown mechanism by which these proteins cross the mycobacterial cell envelope.

microbiology↗

bioRxiv2020

Proteomic and Transcriptomic Analyses of the Hippocampus and Cortex in SUDEP and High-Risk SUDEP Cases

Sudden unexpected death in epilepsy (SUDEP) is the leading type of epilepsy-related death. Severely depressed brain activity in these cases may impair respiration, arousal, and protective reflexes, occurring as a prolonged postictal generalized EEG suppression (PGES) and resulting in a high-risk for SUDEP. In autopsy hippocampus and cortex, we observed no proteomic differences between SUDEP and epilepsy cases, contrasting our previously reported robust differences between epilepsy and controls. Transcriptomics in hippocampus and cortex from surgical epilepsy cases segregated by PGES identified 55 differentially expressed genes (37 protein-coding, 15 lncRNAs, three pending) in hippocampus. Overall, the SUDEP proteome and high-risk SUDEP transcriptome largely reflected other epilepsy cases in the brain regions analyzed, consistent with diverse epilepsy syndromes and comorbidities associated with SUDEP. Thus, studies with larger cohorts and different epilepsy syndromes, as well as additional anatomic regions may identify molecular mechanisms of SUDEP.

neuroscience↗

bioRxiv2020

Proteomic Differences in the Hippocampus and Cortex of Epilepsy Brain Tissue

Epilepsy is a common neurological disorder affecting over 70 million people worldwide, with a high rate of pharmaco-resistance, diverse comorbidities including progressive cognitive and behavioral disorders, and increased mortality from direct (e.g., Sudden Unexpected Death in Epilepsy [SUDEP], accidents, drowning) or indirect effects of seizures and therapies. Extensive research with animal models and human studies provides limited insights into the mechanisms underlying seizures and epileptogenesis, and these have not translated into significant reductions in pharmaco-resistance, morbidities or mortality. To help define changes in molecular signaling networks associated with epilepsy, we examined the proteome of brain samples from epilepsy and control cases. Label-free quantitative mass spectrometry (MS) was performed on the hippocampal CA1-3 region, frontal cortex, and dentate gyrus microdissected from epilepsy and control cases (n=14/group). Epilepsy cases had significant differences in the expression of 777 proteins in the hippocampal CA1-3 region, 296 proteins in the frontal cortex, and 49 proteins in the dentate gyrus in comparison to control cases. Network analysis showed that proteins involved in protein synthesis, mitochondrial function, G-protein signaling, and synaptic plasticity were particularly altered in epilepsy. While protein differences were most pronounced in the hippocampus, similar changes were observed in other brain regions indicating broad proteomic abnormalities in epilepsy. Among the most significantly altered proteins, G-protein Subunit Beta 1 (GNB1) was one of the most significantly decreased proteins in epilepsy in all regions studied, highlighting the importance of G-protein subunit signaling and G-protein-coupled receptors (GPCRs) in epilepsy. Our results provide insights into the molecular mechanisms underlying epilepsy, which may allow for novel targeted therapeutic strategies.

neuroscience↗