Biology subjects

Heskett, M. B.

Publications and source records attributed to Heskett, M. B..

7 recordsLinked to original sources

bioRxiv2025

Autosomal Allelic Inactivation: Variable Replication and Dosage Sensitivity

Autosomal monoallelic gene expression and asynchronous replication between alleles are established features of imprinted genes and genes regulated by allelic exclusion. Inactivation/Stability Centers (I/SCs) are recently described autosomal loci that exhibit epigenetic regulation of allelic expression and replication timing, with differences that can be comparable to those observed between the active and inactive X chromosomes1. Here we characterize >100 autosomal loci with allele-specific epigenetic regulation of replication timing and gene expression, defining them as I/SCs. I/SCs are approximately 1 megabase in size and can contain both protein-coding and noncoding genes. In different single cell derived clones, these genes may be expressed from a single allele, the opposite allele, both alleles, or not expressed at all. This stochastic, yet mitotically stable, pattern indicates that the choice of which allele is expressed is independent of parent of origin and independent of the expression status of the other allele. Similarly, alleles within I/SCs show varying replication timing, either earlier or later, that is also independent of the other allele. Additionally, we identify syntenic loci in the mouse genome that display epigenetic regulation of allelic replication timing, highlighting the genomic organization and conservation of I/SC-associated regulation between human and mouse genomes. The allele-restricted regulation described here creates extensive cellular mosaicism through a stable epigenetic mechanism. This mosaicism impacts numerous dosage-sensitive genes associated with human diseases such as Alzheimer, Parkinson, epilepsy, deafness, and impaired intellectual development.

genomics↗

bioRxiv2025

Myc and Kras cooperate in adult acinar cells to drive phenotypic heterogeneity, metastasis, and therapeutic resistance in a novel pancreatic cancer mouse model

Pancreatic ductal adenocarcinoma (PDAc) is a deadly malignancy, most commonly diagnosed in advanced stages when no curative treatments are available. The development of new models that aid ongoing investigation into the mechanisms by which it initiates, disseminates, and evades treatment is of the utmost importance. In vivo models that accurately recapitulate the features and spectrum of human pancreatic cancer are paramount to make a dent in this disease as two decades of the standard-of-care have failed to substantially improve survival. Here, we take advantage of our finding that post-translational stabiliziation of MYC downstream of the canonical PDAc driver, mutant KRAS, is an early event in PDAc progression to design a novel mouse model of PDAc progression based on deregulated, constituitive expression of Myc and mutant Kras in adult pancreatic acinar cells. Tumors from this KMC model histologically and molecularly recapitulate heterogeneity seen in human PDAc, with a high rate of metastasis to the liver. Cell lines derived from KMC autochthonous PDAc provide new models for orthotopic primary tumors that reliably metastasize to the liver and lung, providing important new tools to efficiently study the metastatic cascade and aid in the develoment of new therapeutics addressing metastatic disease. Cell lines represent distinct molecular subtypes with corresponding differential drug sensitivity. Toghether, this model provides a new and additional tool in the study of pancreatic cancer and the means by which it so deftly evades our best efforts at treatment.

cancer biology↗

bioRxiv2023

Characterization of methylation status of the nuclear hormone receptor DAX-1 in human cancer

The orphan receptor DAX-1 plays an essential role in human development, steroid hormone synthesis and the maintenance of embryonic stem cell pluripotency. Recent studies have demonstrated DAX-1 is involved in cancer development, and, depending on the specific cancer type, has a negative or positive effect on cancer growth. In order to better understand the mechanism of DAX-1 gene regulation in various cancer cell lines, the epigenetic regulation of DAX-1 was investigated. Following confirming levels of DAX-1 expression at both the mRNA and protein levels, the overall methylation status of the DAX-1 gene was probed using methylation-sensitive restriction enzyme analysis. To determine the molecular mechanism of DNA methylation of the DAX-1 gene, chromatin immunoprecipitation assays identified key methylating proteins that localize to specific CpG islands in the DAX-1 promoter. In conclusion, this study demonstrates that methylation of key cytosine residues in CpG islands within the DAX-1 promoter play a central role in regulating DAX-1 expression and varying degrees of methylation result in differences in DAX-1 expression in human cancer cell lines.

molecular biology↗

bioRxiv2022

ASAR RNAs control replication through interactions with dozens of RNA binding proteins

ASARs are a family of very-long noncoding RNAs that control replication timing on individual human autosomes, and are essential for chromosome stability. The eight known ASAR lncRNAs remain closely associated with their parent chromosomes. Analysis of RNA-protein interaction data (from ENCODE) revealed numerous RBPs with significant interactions with multiple ASAR lncRNAs, with several hnRNPs as abundant interactors. An [~]7kb domain within the ASAR6-141 lncRNA shows a striking density of RBP interaction sites. Genetic deletion and ectopic integration assays indicate that this [~]7kb RNA binding protein domain contains functional sequences for controlling replication timing of entire chromosomes in cis. shRNA-mediated depletion of 10 different RNA binding proteins, including HNRNPA1, HNRNPC, HNRNPL, HNRNPM, HNRNPU, or HNRNPUL1, results in dissociation of ASAR lncRNAs from their chromosome territories, and disrupts the synchronous replication that occurs on all autosome pairs, recapitulating the effect of individual ASAR knockouts on a genome-wide scale. Our results further demonstrate the role that ASARs play during the temporal order of genome-wide replication, and we propose that ASARs function as essential RNA scaffolds for the assembly of hnRNP complexes that help maintain the structural integrity of each mammalian chromosome.

genetics↗

bioRxiv2022

Epigenetic Control of Hundreds of Chromosome-Associated lncRNA Genes Essential for Replication and Stability

ASARs are long noncoding RNA genes that control replication timing of entire human chromosomes in cis. The three known ASAR genes are located on human chromosomes 6 and 15, and are essential for chromosome integrity. To identify ASARs on all human chromosomes we utilized a set of distinctive ASAR characteristics that allowed for the identification of hundreds of autosomal loci with epigenetically controlled, allele-restricted behavior in expression and replication timing of coding and noncoding genes, and is distinct from genomic imprinting. Disruption of noncoding RNA genes at five of five tested loci resulted in chromosome-wide delayed replication and chromosomal instability, validating their ASAR activity. In addition to the three known essential cis-acting chromosomal loci, origins, centromeres, and telomeres, we propose that all mammalian chromosomes also contain "Inactivation/Stability Centers" that display allele-restricted epigenetic regulation of protein coding and noncoding ASAR genes that are essential for replication and stability of each chromosome.

genetics↗

bioRxiv2019

Multi-region exome sequencing of ovarian teratomas reveals 2N near-diploid genomes, paucity of somatic mutations, and extensive allelic imbalances shared across mature, immature, and disseminated components

Immature teratoma is a subtype of malignant germ cell tumor of the ovary that occurs most commonly in the first three decades of life, frequently with bilateral ovarian disease. Despite being the second most common malignant germ cell tumor of the ovary, little is known about its genetic underpinnings. Here we performed multi-region whole exome sequencing to interrogate the genetic zygosity, clonal relationship, DNA copy number, and mutational status of 52 pathologically distinct tumor components from 10 females with ovarian immature teratomas, with bilateral tumors present in 5 cases and peritoneal dissemination in 7 cases. We found that ovarian immature teratomas are genetically characterized by 2N near-diploid genomes with extensive loss of heterozygosity and an absence of genes harboring recurrent somatic mutations or known oncogenic variants. All components within a single ovarian tumor (immature teratoma, mature teratoma with different histologic patterns of differentiation, and yolk sac tumor) were found to harbor an identical pattern of loss of heterozygosity across the genome, indicating a shared clonal origin. In contrast, the 4 analyzed bilateral teratomas showed distinct patterns of zygosity changes in the right versus left sided tumors, indicating independent clonal origins. All disseminated teratoma components within the peritoneum (including gliomatosis peritonei) shared a clonal pattern of loss of heterozygosity with either the right or left primary ovarian tumor. The observed genomic loss of heterozygosity patterns indicate that diverse meiotic errors contribute to the formation of ovarian immature teratomas, with 11 out of the 15 genetically distinct clones determined to result from the failure of meiosis I or II. Overall, these findings suggest that copy-neutral loss of heterozygosity resulting from meiotic abnormalities may be sufficient to generate ovarian immature teratomas from germ cells.

cancer biology↗

bioRxiv2019

Reciprocal monoallelic expression of ASAR lncRNA genes controls replication timing of human chromosome 6.

AbstractDNA replication occurs on mammalian chromosomes in a cell-type distinctive temporal order known as the replication timing program. We previously found that disruption of the noncanonical lncRNA genes ASAR6 and ASAR15 results in delayed replication timing and delayed mitotic chromosome condensation of human chromosome 6 and 15, respectively. ASAR6 and ASAR15 display random monoallelic expression, and display asynchronous replication between alleles that is coordinated with other random monoallelic genes on their respective chromosomes. Disruption of the expressed allele, but not the silent allele, of ASAR6 leads to delayed replication, activation of the previously silent alleles of linked monoallelic genes, and structural instability of human chromosome 6. In this report, we describe a second lncRNA gene (ASAR6-141) on human chromosome 6 that when disrupted results in delayed replication timing in cis. ASAR6-141 is subject to random monoallelic expression and asynchronous replication, and is expressed from the opposite chromosome 6 homolog as ASAR6. ASAR6-141 RNA, like ASAR6 and ASAR15 RNAs, contains a high L1 content and remains associated with the chromosome territory where it is transcribed. Three classes of cis-acting elements control proper chromosome function in mammals: origins of replication, centromeres; and telomeres, which are responsible for replication, segregation and stability of all chromosomes. Our work supports a fourth type of essential chromosomal element, \"Inactivation/Stability Centers\", which express ASAR lncRNAs responsible for proper replication timing, monoallelic expression, and structural stability of each chromosome.\n\nAuthor summaryMammalian cells replicate their chromosomes during a highly ordered and cell type-specific program. Genetic studies have identified two long non-coding RNA genes, ASAR6 and ASAR15, as critical regulators of the replication timing program of human chromosomes 6 and 15, respectively. There are several unusual characteristics of the ASAR6 and ASAR15 RNAs that distinguish them from other long non-coding RNAs, including: being very long (>200 kb), lacking splicing of the transcripts, lacking polyadenylation, and being retained in the nucleus on the chromosomes where they are made. ASAR6 and ASAR15 also have the unusual property of being expressed from only one copy of the two genes located on homologous chromosome pairs. Using these unusual characteristics shared between ASAR6 and ASAR15, we have identified a second ASAR lncRNA gene located on human chromosome 6, which we have named ASAR6-141. ASAR6-141 is expressed from the opposite chromosome 6 homolog as ASAR6, and disruption of the expressed allele results in delayed replication of chromosome 6. ASAR6-141 RNA had previously been annotated as vlinc273. The very long intergenic non-coding (vlinc)RNAs represent a recently annotated class of RNAs that are long (>50 kb), non-spliced, and non-polyadenlyated nuclear RNAs. There are currently >2,700 vlincRNAs expressed from every chromosome, are encoded by >15% of the human genome, and with a few exceptions have no known function. Our results suggest the intriguing possibility that the vlinc class of RNAs may be functioning to control the replication timing program of all human chromosomes.

genetics↗