Biology subjects

Mousavi, N.

Publications and source records attributed to Mousavi, N..

9 recordsLinked to original sources

bioRxiv2026

Scalable genotyping in fixed transcriptomes resolves clonal heterogeneity via single-cell sequencing

Despite the promise of single-cell transcriptomics for understanding cell states in heterogeneous populations, widely used platforms have limited ability to link transcriptional states to somatic mutations within the same cells. Here, we introduce Genotyping in Fixed Transcriptomes (GIFT) for the simultaneous detection of large numbers of targeted genetic variants with whole transcriptome profiles in single cells. The core innovation of GIFT is a rationally designed gapfilling reaction between adjacent single-stranded DNA (ssDNA) probes that barcodes native transcript sequence to enable highly-specific targeted mutation detection. GIFT achieves greater than 99% genotyping accuracy and flexible capture of hundreds of mutations per cell, including in formalin-fixed, paraffin-embedded (FFPE) tissue, enabling clonal lineage tracing in heterogeneous settings. We demonstrate the unique scalability of GIFT by profiling more than 700,000 cells from 35 donors with myeloproliferative neoplasms (MPN), revealing mutation-dependent hematopoietic responses to systemic inflammation associated with the characteristic JAK2V617 mutation, including an allelic dose gradient of interferon-associated transcriptional programs and priming of hematopoietic stem cells that develop into divergent disease states. The technical advantages of GIFT enable direct resolution of genotype-to-phenotype relationships via clonal tracing with comprehensive cell-state measurements at single-cell resolution.

genomics↗

bioRxiv2024

Topography of putative bidirectional interaction between hippocampal sharp wave ripples and neocortical slow oscillations

Systems consolidation relies on coordination between hippocampal sharp-wave ripples (SWRs) and neocortical UP/DOWN states during sleep. However, whether this coupling exists across neocortex and the mechanisms enabling it remain unknown. By combining electrophysiology in mouse hippocampus (HPC) and retrosplenial cortex (RSC) with widefield imaging of dorsal neocortex, we found spatially and temporally precise bidirectional hippocampo-neocortical interaction. HPC multi-unit activity and SWR probability was correlated with UP/DOWN states in mouse default mode network, with highest modulation by RSC in deep sleep. Further, some SWRs were preceded by the high rebound excitation accompanying DMN DOWN[->]UP transitions, while large-amplitude SWRs were often followed by DOWN states originating in RSC. We explain these electrophysiological results with a model in which HPC and RSC are weakly coupled excitable systems capable of bi-directional perturbation and suggest RSC may act as a gateway through which SWRs can perturb downstream cortical regions via cortico-cortical propagation of DOWN states.

neuroscience↗

bioRxiv2024

P3 site-directed mutagenesis: An efficient method based on primer pairs with 3-prime overhangs

Site-directed mutagenesis is a fundamental tool indispensable for protein and plasmid engineering. An important technological question is how to achieve the efficiency at the ideal level of 100%. Based on complementary primer pairs, the QuickChange method has been widely used, but it requires significant improvements due to its low efficiency and frequent unwanted mutations. An alternative and innovative strategy is to utilize primer pairs with 3-prime overhangs, but this approach has not been fully developed. As the first step towards reaching the efficiency of 100%, we have optimized this approach systematically (such as use of newly designed short primers, test of different Pfu DNA polymerases and modification of PCR parameters) and evaluated the resulting method extensively with over 100 mutations on 12 mammalian expression vectors, ranging from 7.0-13.4 kb in size and encoding ten epigenetic regulators with links to cancer and neurodevelopmental disorders. We have also tested the new method with two expression vectors for the SARS-COV-2 spike protein. Compared to the QuickChange method, the success rate has increased substantially, with an average efficiency of [~]50%, with some at or close to 100 percent, and requiring much less time for engineering various mutations. Therefore, we have developed a new site-directed mutagenesis method for efficient and economical generation of various mutations. Notably, the method failed with a human KAT2B expression plasmid that possesses extremely GC-rich sequences. Thus, this study also sheds light on how to improve the method for developing ideal mutagenesis methods with the efficiency at [~]100% for a wide spectrum of plasmids.

molecular biology↗

bioRxiv2023

A deep population reference panel of tandem repeat variation

Tandem repeats (TRs) represent one of the largest sources of genetic variation in humans and are implicated in a range of phenotypes. Here we present a deep characterization of TR variation based on high coverage whole genome sequencing from 3,550 diverse individuals from the 1000 Genomes Project and H3Africa cohorts. We develop a method, EnsembleTR, to integrate genotypes from four separate methods resulting in high-quality genotypes at more than 1.7 million TR loci. Our catalog reveals novel sequence features influencing TR heterozygosity, identifies population-specific trinucleotide expansions, and finds hundreds of novel eQTL signals. Finally, we generate a phased haplotype panel which can be used to impute most TRs from nearby single nucleotide polymorphisms (SNPs) with high accuracy. Overall, the TR genotypes and reference haplotype panel generated here will serve as valuable resources for future genome-wide and population-wide studies of TRs and their role in human phenotypes.

genomics↗

bioRxiv2022

Genome-wide selection inference at short tandem repeats

Short tandem repeats (STRs) comprising repeated sequences of 1-6 bp are one of the largest sources of genetic variation in humans. STRs are known to contribute to a variety of disorders, including Mendelian diseases, complex traits, and cancer. Based on their functional importance, mutations at some STRs are likely to introduce negative effects on reproductive fitness over evolutionary time. We previously developed SISTR (Selection Inference at STRs), a population genetics framework to measure negative selection against individual STR alleles. Here, we extend SISTR to enable joint estimation of the distribution of selection coefficients across a set of STRs. This method (SISTR2) allows for more accurate analysis of a broader range of STRs, including loci with low mutation rates. We apply SISTR2 to explore the range of feasible mutation parameters and demonstrate substantial variation in mutation and selection parameters across different classes of STRs. Finally, we show that de novo STR mutations tend to confer a greater selective burden compared to standing STR variation in the population and measure the relative burden of STRs vs. single nucleotide variants in a typical genome. Overall, we anticipate that the evolutionary insights gained from this study will be important for future studies of variation at STRs and their role in evolution and disease.

genomics↗

bioRxiv2022

A novel quantitative trait locus implicates Msh3 in the propensity for genome-wide short tandem repeat expansions in mice

Short tandem repeats (STRs) are a class of rapidly mutating genetic elements characterized by repeated units of 1 or more nucleotides. We leveraged whole genome sequencing data for 152 recombinant inbred (RI) strains from the BXD family derived from C57BL/6J and DBA/2J mice to study the effects of genetic background on genome-wide patterns of new mutations at STRs. We defined quantitative phenotypes describing the numbers and types of germline STR mutations in each strain and identified a locus on chromosome 13 associated with the propensity of STRs to expand. Several dozen genes lie in the QTL region, including Msh3, a known modifier of STR stability at pathogenic repeat expansions in mice and humans. Detailed analysis of the locus revealed a cluster of variants near the 5 end of Msh3, including multiple protein-coding variants within the DNA mismatch recognition domain of MSH3, and a retrotransposon insertion overlapping an annotated exon. Additionally, gene expression analysis demonstrates co-localization of this QTL with expression QTLs for multiple nearby genes, including Msh3. Our results suggest a novel role for Msh3 in regulating genome-wide patterns of germline STR mutations and demonstrate that inherited genetic variation can contribute to variability in accumulation of new mutations across individuals.

genomics↗

bioRxiv2020

TRTools: a toolkit for genome-wide analysis of tandem repeats

SummaryA rich set of tools have recently been developed for performing genome-wide genotyping of tandem repeats (TRs). However, standardized tools for downstream analysis of these results are lacking. To facilitate TR analysis applications, we present TRTools, a Python library and a suite of command-line tools for filtering, merging, and quality control of TR genotype files. TRTools utilizes an internal harmonization module making it compatible with outputs from a wide range of TR genotypers. AvailabilityTRTools is freely available at https://github.com/gymreklab/TRTools. Contactmgymrek@eng.ucsd.edu Supplementary informationSupplementary data are available at bioRxiv.

bioinformatics↗

bioRxiv2020

The contribution of de novo tandem repeat mutations to autism spectrum disorders

Autism Spectrum Disorder (ASD) is an early onset developmental disorder characterized by deficits in communication and social interaction and restrictive or repetitive behaviors1,2. Family studies demonstrate that ASD has a significant genetic basis3 with contributions both from inherited and de novo variants. While the majority of variance in liability to ASD is estimated to arise from common genetic variation4, it has been estimated that de novo mutations may contribute to 30% of all simplex cases, in which only a single child is affected per family5. Tandem repeats (TRs), consisting of approximately 1-20bp motifs repeated in tandem, comprise one of the largest sources of de novo mutations in humans6. Yet, largely due to technical challenges they present, de novo TR mutations have not yet been characterized on a genome-wide scale, and their contribution to ASD remains unexplored. Here, we develop novel bioinformatics tools for identifying and prioritizing de novo TR mutations from whole genome sequencing (WGS) data and use these to perform a genome-wide characterization of de novo TR mutations in ASD-affected probands and unaffected siblings. Compared to recent work on TRs in ASD7, we explicitly infer mutation events and their precise changes in repeat copy number, and primarily focus on more prevalent stepwise copy number changes rather than large or complex expansions. Our results demonstrate a significant genome-wide excess of TR mutations in ASD probands. TR mutations in probands tend to be larger, enriched in fetal brain regulatory regions, and predicted to be more evolutionarily deleterious compared to mutations observed in unaffected siblings. Overall, our results highlight the importance of considering repeat variants in future studies of de novo mutations.

genomics↗

bioRxiv2018

Profiling the genome-wide landscape of tandem repeat expansions

Tandem Repeat (TR) expansions have been implicated in dozens of genetic diseases, including Huntingtons Disease, Fragile X Syndrome, and hereditary ataxias. Furthermore, TRs have recently been implicated in a range of complex traits, including gene expression and cancer risk. While the human genome harbors hundreds of thousands of TRs, analysis of TR expansions has been mainly limited to known pathogenic loci. A major challenge is that expanded repeats are beyond the read length of most next-generation sequencing (NGS) datasets. We present GangSTR, a novel algorithm for genome-wide profiling of both normal and expanded TRs. GangSTR extracts information from paired-end reads into a unified model to estimate maximum likelihood TR lengths. We validated GangSTR on real and simulated TR expansions and show that GangSTR outperforms alternative methods. We applied GangSTR to more than 150 individuals to profile the landscape of TR expansions in a healthy population and validated novel expansions using orthogonal technologies. Our analysis revealed that each individual harbors dozens of TR alleles longer than standard read lengths and identified hundreds of potentially mis-annotated TRs in the reference genome. GangSTR is packaged as a standalone tool that will likely enable discovery of novel pathogenic variants not currently accessible from NGS.

genomics↗