Biology subjects

Maguire, K. M.

Publications and source records attributed to Maguire, K. M..

3 recordsLinked to original sources

bioRxiv2026

Multi-Modal Kinome Profiling Discovers Mesenchymal-Like Polarity Networks that Underly Directed Hepatocellular Carcinoma Cell Migration

Metastasis and associated therapy resistance remain the principal drivers of cancer related death, and there is a pressing need for a deeper mechanistic understanding and anti-metastatic therapies. For patients that suffer from hepatocellular carcinomas (HCCs), which are the most common primary liver cancers, frequent systemic metastasis results in bleak 5-year survival prognoses of only 4%. To metastasize, carcinoma cells must acquire an invasive phenotype, which typically requires switching from an epithelial-like apical-basal polarity to the front-rear polarity of mesenchymal-like cells. Signaling cues that originate in the tumor microenvironment can activate cellular morphogenic programs that drive polarity switching, like the epithelial-mesenchymal transition (EMT). Protein kinases control most cell signaling pathways and are highly actionable drug targets; however, systematic studies determining the kinases that underly the epithelial-mesenchymal polarity switch (EMPS) are lacking. We developed an assay platform that integrates mass spectrometry (MS)-based kinome profiling, broadly capturing kinase network activity, with chemical genetic screening using selective kinase inhibitors and quantitative phase imaging (QPI), serving as the phenotypic readout. Applying this approach that we dubbed morphokin-MS, to epithelial-like HCC cell lines that we induced to undergo EMPS identified a conserved network of 12 kinases that contributed to HCC cell polarity switching and directed cell migration; MS-based kinome profiling of 17 HCC patient tumors showed that these kinase are frequently upregulated in human tumors. morphokin-MS also revealed that death-associated protein kinase 3 (DAPK3) is one of the principal drivers of the EMPS and directed HCC cell migration. Thus, our mechanistic studies revealed that DAPK3 forms a complex with DAPK1 and filamin-A inter-acting protein 1-like (FILIP1L), which act as scaffold proteins that recruit DAPK3 to the centrosome. Pharmaco-logical and genetic inhibition of the DAPK1-DAPK3-FILIP1L complex blocked centrosome repositioning and microtubule polarization toward the leading edge of mesenchymal-like HCC cells, directed cell migration, and invasion. Our morphokin-MS method and comprehensive kinome profiling data will serve as a valuable resource for the cancer research community; our discovery of an inducible mesenchymal-like DAPK1-DAPK3-FILIP1L polarity complex that controls centrosome positioning in motile HCC cells may lead to the development of novel therapeutics for combatting cancer metastasis.

cancer biology↗

bioRxiv2024

diaPASEF-Powered Chemoproteomics Enables Deep Kinome Interaction Profiling

Protein-protein interactions (PPIs) underlie most biological functions. Devastating human conditions like cancers, neurological disorders, and infections, hijack PPI networks to initiate disease, and to drive disease progression. Understanding precisely how diseases remodel PPI networks can, therefore, help clarify disease mechanisms and identify therapeutic targets. Protein kinases control most cellular processes through protein phosphorylation. The 518 human kinases, known as the kinome, are frequently dysregulated in disease and highly druggable with ATP-competitive inhibitors. Kinase activity, localization, and substrate recognition are regulated by dynamic PPI networks composed of scaffolding and adapter proteins, other signaling enzymes like small GTPases and E3 ligases, and phospho-substrates. Accordingly, mapping kinase PPI networks can help determine kinome activation states, and, in turn, cellular activation states; this information can be used for studying kinase-mediated cell signaling, and for prioritizing kinases for drug discovery. Previously, we have developed a high-throughput method for kinome PPI mapping based on mass spectrometry (MS)-based chemoproteomics that we named kinobead competition and correlation analysis (kiCCA). Here, we introduce 2nd generation (gen) kiCCA which utilizes data-independent acquisition (dia) with parallel accumulation serial fragmentation (PASEF) MS and a re-designed CCA algorithm with improved selection criteria and the ability to predict multiple kinase interaction partners of the same proteins. Using neuroblastoma cell line models of the noradrenergic-mesenchymal transition (NMT), we demonstrate that 2nd gen kiCCA (1) identified 6.1-times more kinase PPIs in native cell extracts compared to our 1st gen approach, (2) determined kinase-mediated signaling pathways that underly the neuroblastoma NMT, and (3) accurately predicted pharmacological targets for manipulating NMT states. Our 2nd gen kiCCA method is broadly useful for cell signaling research and kinase drug discovery.

biochemistry↗

bioRxiv2024

HAF Prevents Hepatocyte Apoptosis and Hepatocellular Carcinoma through Transcriptional Regulation of the NF-κB pathway.

BackgroundHepatocellular carcinoma (HCC) incidence is increasing worldwide due to the obesity epidemic, which drives metabolic dysfunction-associated steatohepatitis (MASH) that can lead to HCC. However, the molecular pathways that lead to MASH-HCC are poorly understood. We have previously reported that male mice with global haploinsufficiency of hypoxia-associated factor, HAF (SART1+/-) spontaneously develop MASH/HCC. However, the cell type(s) responsible for HCC associated with HAF loss are unclear. ResultsSART1-floxed mice were crossed with mice expressing Cre-recombinase within hepatocytes (Alb-Cre; hepS-/-) or macrophages (LysM-Cre, macS-/-). Only hepS-/- mice (both male and female) developed HCC suggesting that HAF protects against HCC primarily within hepatocytes. HAF-deficient macrophages showed decreased P-p65 and P-p50 and in many major components of the NF-{kappa}B pathway, which was recapitulated using HAF siRNA in vitro. HAF depletion increased apoptosis both in vitro and in vivo, suggesting that HAF mediates a tumor suppressor role by suppressing hepatocyte apoptosis. We show that HAF regulates NF-{kappa}B activity by controlling transcription of TRADD and RIPK1. Mice fed a high-fat diet (HFD) showed marked suppression of HAF, P-p65 and TRADD within their livers after 26 weeks, but manifest profound upregulation of HAF, P-65 and TRADD within their livers after 40 weeks of HFD, implicating deregulation of the HAF-NF-{kappa}B axis in the progression to MASH. In humans, HAF was significantly decreased in livers with simple steatosis but significantly increased in HCC compared to normal liver. ConclusionsHAF is novel transcriptional regulator of the NF-{kappa}B pathway that protects against hepatocyte apoptosis and is a key determinant of cell fate during progression to MASH and MASH-HCC.

cancer biology↗