Project description:Human leukocyte antigen loss of heterozygosity (HLA LOH) allows cancer cells to escape immune recognition by deleting HLA alleles, causing the suppressed presentation of tumor neoantigens. Despite its importance in immunotherapy response, few methods exist to detect HLA LOH, and their accuracy is not well understood. Here, we develop DASH (Deletion of Allele-Specific HLAs), a machine learning-based algorithm to detect HLA LOH from paired tumor-normal sequencing data. With cell line mixtures, we demonstrate increased sensitivity compared to previously published tools. Moreover, our patient-specific digital PCR validation approach provides a sensitive, robust orthogonal approach that could be used for clinical validation. Using DASH on 610 patients across 15 tumor types, we find that 18% of patients have HLA LOH. Moreover, we show inflated HLA LOH rates compared to genome-wide LOH and correlations between CD274 (encodes PD-L1) expression and microsatellite instability status, suggesting the HLA LOH is a key immune resistance strategy.
Project description:Early B cell factor 1 (EBF1) is one of the key transcription factors required for orchestrating B-cell lineage development. Although studies have shown that Ebf1 haploinsufficiency is involved in the development of leukemia, no study has been conducted that characterizes the global effect of Ebf1 heterozygosity on the proteome of pro-B lymphocytes. Here, we employ both DIA (Data Independent Acquisition) and shotgun DDA (Data Dependent Acquisition) workflows for profiling proteins that are differently expressed between Ebf1+/+ and Ebf1+/- cells. Both DDA and DIA were able to reveal the downregulation of the EBF1 transcription factor in Ebf1+/- pro-B lymphocytes. Further examination of differentially expressed proteins by DIA revealed that, similar to EBF1, the expression of other B-cell lineage regulators, such as TCF3 and Pax5, is also down-regulated in Ebf1 heterozygous cells. Functional DIA analysis of differentially expressed proteins showed that EBF1 heterozygosity resulted in the deregulation of at least 8 transcription factors involved in lymphopoiesis, and to the deregulation of key proteins playing crucial roles in survival, development and differentiation of pro-B lymphocytes.
Project description:To study the relationship between loss of heterozygosity (LOH) at the human leukocyte antigen (HLA) locus and the pathogenicity and clinicopathological features of thymic epithelial tumors (TET).Tumor and adjacent normal tissues were isolated from 36 TET patients. Five microsatellite loci (D6S1666, D6S265, D6S273, DS6276, and D6S291) within the HLA locus were amplified by polymerase chain reaction. DNA sequencing was used to measure the frequency of microsatellite LOH.LOH was identified in at least one locus in 83.6% of TET patients. LOH frequency at D6S1666, D6S265, D6S273, D6S276, and D6S291 was 44.4%, 16.7%, 30.5%, 38.9%, and 36.1% respectively. There was no significant association between LOH frequency in TET with tumor severity, or in the presence or absence of myasthenia gravis.D6S1666, D6S265, D6S273, DS6S276, and D6S29 are sensitive loci for studying microsatellite LOH in TET. LOH within the HLA complex is implicated in the occurrence and development of TET, with the HLA-DQA1 gene likely involved. However, an understanding of the relationship between LOH and the clinicopathological features of TET requires a larger sample size than that of the present study.
Project description:Genome-wide methylation patterns observed in primary human leukocyte subsets were used to identify cell-typeM-^Vspecific regulatory hypomethylated regions. To investigate the possible effect of these regions on gene expression, a transcriptomics analysis was performed with mRNA isolated from the leukocyte subsets.
Project description:CGH arrays for Smukowski Heil, et al MBE 2017. Hybridization is often considered maladaptive, but sometimes hybrids can invade new ecological niches and adapt to novel or stressful environments better than their parents. The genomic changes that occur following hybridization that facilitate genome resolution and/or adaptation are not well understood. Here, we address these questions using experimental evolution of de novo interspecific hybrid yeast Saccharomyces cerevisiae x Saccharomyces uvarum and their parentals. We evolved these strains in nutrient limited conditions for hundreds of generations and sequenced the resulting cultures to identify genomic changes. Analysis of 16 hybrid clones and 16 parental clones identified numerous point mutations, copy number changes, and loss of heterozygosity events, including species biased amplification of nutrient transporters. We focused on a particularly interesting example, in which we saw repeated loss of heterozygosity at the high affinity phosphate transporter gene PHO84 in both intra- and interspecific hybrids. Using allele replacement methods, we tested the fitness of different alleles in hybrid and S. cerevisiae strain backgrounds and found that the loss of heterozygosity is indeed the result of selection on one allele over the other in both S. cerevisiae and the hybrids. This is an example where hybrid genome resolution is driven by positive selection on existing heterozygosity, and demonstrates that even infrequent outcrossing may have lasting impacts on adaptation.