Biology subjects

Nefzger, C. M.

Publications and source records attributed to Nefzger, C. M..

6 recordsLinked to original sources

bioRxiv2024

MMCCI: Multimodal Cell-Cell Interaction Integrative Analysis of Single Cell and Spatial Data

Cell-cell interaction (CCI) analyses are an indispensable tool for harnessing the detail and depth of spatial and single-cell transcriptomics datasets by inferring inter-cellular communications, but no methods to integrate CCI results across samples exist currently. To address this, we have developed a computational pipeline, Multimodal CCI (MMCCI), to statistically integrate and analyze CCI results from existing popular CCI tools. We benchmarked MMCCIs integration on single-cell spatial datasets and found it to be highly accurate compared to simpler methods. We utilized MMCCIs integration and downstream biological analyses to uncover global and differential interaction patterns in multimodal aging brain and melanoma spatial datasets.

bioinformatics↗

bioRxiv2022

Organisation of gene programs revealed by unsupervised analysis of diverse gene-trait associations

Genome wide association studies provide statistical measures of gene-trait associations that reveal how genetic variation influences phenotypes. This study develops an unsupervised dimensionality reduction method called UnTANGLeD (Unsupervised Trait Analysis of Networks from Gene Level Data) which organises 16,849 genes into discrete gene programs by measuring the statistical association between genetic variants and 1,393 diverse complex traits. UnTANGLeD reveals 173 gene clusters enriched for protein-protein interactions and highly distinct biological processes governing development, signalling, disease, and homeostasis. We identify diverse gene networks with robust interactions but not associated with known biological processes. Analysis of independent disease traits shows that UnTANGLeD gene clusters are conserved across all complex traits, providing a simple and powerful framework to predict novel gene candidates and programs influencing orthogonal disease phenotypes. Collectively, this study demonstrates that gene programs co-ordinately orchestrating cell functions can be identified without reliance on prior knowledge, providing a method for use in functional annotation, hypothesis generation, machine learning and prediction algorithms, and the interpretation of diverse genomic data.

genomics↗

bioRxiv2022

Retrotransposon instability dominates the acquired mutation landscape of mouse induced pluripotent stem cells

Induced pluripotent stem cells (iPSCs) can in principle differentiate into any cell of the body, and have revolutionized biomedical research and regenerative medicine. Unlike their human counterparts, mouse iPSCs (miPSCs) are reported to silence transposable elements (TEs) and prevent TE-mediated mutagenesis. Here we applied short-read or Oxford Nanopore Technologies (ONT) long-read genome sequencing to 38 bulk miPSC lines reprogrammed from 10 parental cell types, and 18 single-cell miPSC clones. While single nucleotide variants and structural variants restricted to miPSCs were rare, we found 83 de novo TE insertions, including examples intronic to Brca1 and Dmd. LINE-1 (L1) retrotransposons were profoundly hypomethylated in miPSCs, beyond other TEs and the genome overall, and harbored alternative protein-coding gene promoters. Treatment with the L1 inhibitor lamivudine did not hinder reprogramming and efficiently blocked endogenous retrotransposition, as detected by ONT sequencing. These experiments reveal the complete spectrum and potential significance of mutations acquired by miPSCs.

genomics↗

bioRxiv2020

TINC - a method to dissect transcriptional complexes at single locus resolution - reveals novel Nanog regulators in mouse embryonic stem cells

Cellular identity is ultimately controlled by transcription factors (TFs), which bind to specific regulatory elements (REs) within the genome to regulate gene expression and cell fate changes. While recent advances in genome-wide epigenetic profiling techniques have significantly increased our understanding of which REs are utilized in which cell type, it remains largely unknown which TFs and cofactors interact with these REs to modulate gene expression. A major hurdle in dissecting the whole composition of a multi-protein complex formed at a specific RE is the shortage of appropriate techniques. We have developed a novel method termed TALE-mediated Isolation of Nuclear Chromatin (TINC). TINC utilizes epitope-tagged TALEs to isolate a specific genomic region from the mammalian genome and includes a nuclei isolation and chromatin enrichment step for increased specificity. Upon cross-linking of the cells and isolation of the chromatin, the target region is purified based on affinity purification of the TALE and associated nucleic acid and protein molecules can be subjected to further analyses. A key TF in the pluripotency network and therefore in embryonic stem cells (ESCs) is NANOG. It is currently not fully understood how Nanog expression is regulated and consequently it remains unclear how the ESC state is maintained. Using TINC we dissected the protein complex formed at the Nanog promoter in mouse ESCs and identified many known and numerous novel factors.

genomics↗

bioRxiv2019

Xmas ESC: A new female embryonic stem cell system that reveals the BAF complex as a key regulator of the establishment of X chromosome inactivation

Although female pluripotency significantly differs to male, complications with in vitro culture of female embryonic stem cells (ESC) have severely limited the use and study of these cells. We report a replenishable female ESC system, Xmas, that has enabled us to optimise a protocol for preserving the XX karyotype. Our protocol also improves male ESC fitness. We utilised our Xmas ESC system to screen for regulators of the female-specific process of X chromosome inactivation, revealing chromatin remodellers Smarcc1 and Smarca4 as key regulators of establishment of X inactivation. The remodellers create a nucleosome depleted region at gene promotors on the inactive X during exit from pluripotency, without which gene silencing fails. Our female ESC system provides a tractable model for XX ESC culture that will expedite study of female pluripotency and has enabled us to discover new features of the female-specific process of X inactivation.

cell biology↗

bioRxiv2018

Generation of four iPSC lines from peripheral blood mononuclear cells (PBMCs) of an Attention Deficit Hyperactivity Disorder (ADHD) individual and a healthy sibling in an Australia-Caucasian family

Peripheral blood mononuclear cells were donated by a male teenager with clinically diagnosed Attention Deficit Hyperactivity Disorder under the Diagnostic and Statistical Manual of Mental Disorders IV criteria and his unaffected male sibling. Induced pluripotent stem cells were developed using integration-free Sendai Reprogramming factors containing OCT, SOX2, KLF4, and c-MYC. All four iPSC lines displayed pluripotent cell morphology, pluripotency-associated factors at protein level, alkaline phosphatase enzymatic activity, male karyotype of 46, XY, and in vitro differentiation capacity into all the three germ layers and negative for Mycoplasma.

neuroscience↗