Biology subjects

Barlow, J. H.

Publications and source records attributed to Barlow, J. H..

4 recordsLinked to original sources

bioRxiv2026

Integrative proteomic analysis and molecular dynamics simulations of ANKLE2 reveal mechanisms of microcephaly

ANKLE2 is a scaffolding protein with crucial roles in neuroprogenitor cell division and embryonic brain development. Pathogenic variants in ANKLE2 cause primary microcephaly, a congenital disorder characterized by impaired neurodevelopment. Nonetheless, the underlying dysfunction of ANKLE2 is not understood. Here, we define the ANKLE2 protein interaction landscape, which includes cell division proteins and novel microcephaly candidates in the APC (anaphase-promoting complex). Analysis of six pathogenic variants reveals changes in this interactome that likely result in microcephaly. Using molecular simulations, we identify the structural consequences of five pathogenic substitutions, including disruption of important helical structures. Combined, these techniques suggest loss of interaction with the PP2A (protein phosphatase 2A) complex is a common mechanism for ANKLE2 pathogenesis. Phosphoproteomics reveals widespread alterations in ANKLE2- and PP2A-dependent phosphorylation of cell division proteins. Together, our integrative proteomics and simulation approach improves the molecular understanding of ANKLE2 function and its pathogenic changes in primary microcephaly.

systems biology↗

bioRxiv2026

Reduced B cell activation in Germinal center reaction of the mouse model of Li-Fraumeni syndrome

The TP53 mutation associated with Li-Fraumeni syndrome in humans is known to exhibit p53 gain-of-function properties leading to early cancer onset. To explore whether these patients also have compromised immune responses due to p53 mutations, we investigated the humoral immune response using a Li-Fraumeni mouse model harboring a structural mutation, Trp53R172H (equivalent to codon 175 in humans). Trp53R172H mice were immunized with sheep red blood cells, and the Germinal center response was monitored. Our results revealed that SRBC-immunized Trp53R172H mice exhibit reduced B cell activation during the GC reaction. These suggest a selective role for p53 in promoting B cell activation early in the GC reaction prior to BCL6 upregulation. We propose that impaired B cell activation in Li-Fraumeni patients could contribute to immune deficiencies and heightened susceptibility to autoimmune disorders, potentially influencing the development of secondary cancers and impairing therapeutic responses to chemotherapies and immunotherapies. Key PointsO_LITrp52R172H mice exhibit reduced B cell activation during the Germinal center reaction C_LIO_LIRatio of activated B cells in DZ to LZ remain unchanged in Trp52R172H mice. C_LI

cancer biology↗

bioRxiv2024

Stress Induced Signaling Pathways in Burkitt Lymphoma Play Novel Mechanisms in Fate Determination and Pathogenesis of Germinal Center-Derived B-Lymphomas

B cell receptor signaling, NF-{kappa}B signaling, BCL6 and p53 regulation are essential for germinal center (GC) B cell fate. Dysregulation of these pathways drives the pathogenesis and treatment resistance of GC-derived B-lymphomas (GCDBL). To explore how these pathways affect GCDBL fate and pathogenesis, we treated Raji cells (a GCDBL and Burkitts lymphoma) with mild hydroxyurea (HU) to simulate genotoxic stress encountered by GC B cells. Genome-wide mapping of histone H3K4me3 and p53 target analysis in HU-treated Raji cells combined with transcriptome analysis of human tonsil GC B cells identified ATAD2B (a p53 target) as differentially expressed. We found that p53 suppresses ATAD2B and ATAD2, while ATAD2 and BCL6 transcripts positively correlate in DLBCLs, suggesting that p53 regulates BCL6 in GC B cells via ATAD2 suppression. We propose that p53 regulation of BCL6 quality assures GC B cells before GC exit. Unlike BCL6 suppression of IFN-{gamma} and NF-{kappa}B signaling in GC B cells, we identified IFNGR1 as a loosely bound BCL6 target and observed loss of BCL6 regulation on genes encoding inhibitory subunits of NF-{kappa}B signaling in B-lymphoma treated with a Bruton tyrosine kinase (BTK) inhibitor. These adaptations, alongside with prevalent genetic inactivation of NF-{kappa}B inhibitory genes in DLBLCs, likely contribute to DLBCL pathogenesis and therapy resistance. Our findings highlight the pivotal role of the p53-BCL6 axis in GC B cell fate and its dysregulation in DLBCL pathogenesis and chemoresistance. Key HighlightsO_LIGenes induced in human Burkitts lymphoma under genotoxic stress are largely independent of histone H3K4me3 marks at transcriptional start sites (TSS). C_LIO_LILoss of BCL6 regulation on genes encoding components of IFN-{gamma} signaling is associated with reduced survival and the pathogenesis of DLBCL. C_LIO_LIInactivating mutations in genes encoding components of the NF-{kappa}B inhibitory subunit serve as an adaptation for DLBCL pathogenesis. C_LIO_LIBCL6 expression is correlated with ATAD2 overexpression in DLBCL and solid tumors. C_LI

cancer biology↗

bioRxiv2021

Genomic patterns of transcription-replication interactions in mouse primary B cells

Conflicts between transcription and replication machinery are a potent source of replication stress and genome stability; however, no technique currently exists to identify endogenous genomic locations prone to transcription-replication interactions. Here, we report a novel method to identify genomic loci prone to transcription-replication interactions termed transcription-replication immunoprecipitation on nascent DNA sequencing, TRIPn-Seq. TRIPn-Seq employs the sequential immunoprecipitation of RNA polymerase 2 phosphorylated at serine 5 (RNAP2s5) followed by enrichment of nascent DNA previously labeled with bromodeoxyuridine. Using TRIPn-Seq, we mapped 1,009 unique transcription-replication interactions (TRIs) in mouse primary B cells characterized by a bimodal pattern of RNAP2s5, bidirectional transcription, an enrichment of RNA:DNA hybrids, and a high probability of forming G-quadruplexes. While TRIs themselves map to early replicating regions, they exhibit enhanced Replication Protein A association and replication fork termination, marks of replication stress. TRIs colocalize with double-strand DNA breaks, are enriched for deletions, and accumulate mutations in tumors. We propose that replication stress at TRIs induces mutations potentially contributing to age-related disease, as well as tumor formation and development.

genomics↗