Biology subjects

Godoy, P. M.

Publications and source records attributed to Godoy, P. M..

5 recordsLinked to original sources

bioRxiv2022

Cross-species analysis reveals unique and shared roles of Sox9 and Sox10 (SOXE family) transcription factors in melanoma

SOX9 and SOX10 are two highly similar transcription factors with nearly 100% identity at their DNA binding domains. Both transcription factors play key but distinct roles in neural crest cell fate specification and melanoma formation. High expression of SOX9 and SOX10 appear to be mutually exclusive, with high SOX10 characteristic of proliferative melanoma and high SOX9 characteristic of metastatic melanoma. To further elucidate the role of SOX9 in melanoma, we over-express SOX9 in a zebrafish melanoma model and a human melanoma cell line. Analysis of tumor onset, binding dynamics, and transcriptional identities supports the notion of SOX9 driving a more mesenchymal signature, which is important for metastasis. Additionally, we identified a potential mechanism of SOX9 down-regulation via analysis of a functional and recurrent non-coding variant in human melanoma. Altogether, our results present a dosage-dependent role of SOX9 and, likely, SOX10 in the melanoma lifespan.

cancer biology↗

bioRxiv2022

Functional analysis of recurrent non-coding variants in human melanoma

Small nucleotide variants in non-coding regions of the genome can alter transcriptional regulation, leading to changes in gene expression which can activate oncogenic gene regulatory networks. Melanoma is heavily burdened by non-coding variants, representing over 99% of total genetic variation, including the well-characterized TERT promoter mutation. However, the compendium of regulatory non-coding variants is likely still functionally under-characterized. We developed a pipeline to identify hotspots, i.e. recurrently mutated regions, in melanoma containing putatively functional non-coding somatic variants that are located within predicted melanoma-specific regulatory regions. We identified hundreds of statistically significant hotspots, including the hotspot containing the TERT promoter variants, and focused in on a hotspot in the promoter of CDC20. We found that variants in the promoter of CDC20, which putatively disrupt an ETS motif, lead to lower transcriptional activity in reporter assays. Using CRISPR/Cas9, we generated an indel in the CDC20 promoter in a human A375 melanoma cell line and observed decreased expression of CDC20, changes in migration capabilities, and an altered transcriptional state previously associated with neural crest transcriptional programs and melanoma initiation. Overall, our analysis prioritized several recurrent functional non-coding variants that, through downregulation of CDC20, led to perturbation of key melanoma phenotypes.

genomics↗

bioRxiv2019

Comparison of miRNA profiling methods using synthetic miRNA pools and standardized exRNA samples reveals substantial performance differences

MicroRNAs (miRNAs) found in biofluids play functional roles in health and in disease pathogenesis, underpinning their potential as clinical biomarkers. Several platforms for measurement of extracellular RNAs have recently become available. We evaluated the reproducibility, accuracy, sensitivity, and specificity of four miRNA quantification platforms, including one widely used discovery approach (small RNA-seq) and three targeted platforms (FirePlex, EdgeSeq, and nCounter). Using pools of synthetic miRNAs, we observed that reproducibility was highest for RNA-seq and EdgeSeq, that all three targeted platforms had lower bias than RNA-seq, and that RNA-seq had the best ability to distinguish between present and absent sequences. Overall reproducibility was lower for plasma samples than synthetic miRNA pools. We compared expression of placental miRNAs in plasma from pregnant and non-pregnant women and observed expected differences with RNA-seq and EdgeSeq, but not FirePlex or nCounter. We conclude that differences in performance among miRNA profiling platforms impact their relative utility as potential assay systems for clinical biomarkers.

genomics↗

bioRxiv2018

Large differences in small RNA composition between human biofluids

Extracellular miRNAs and other small RNAs are implicated in cellular communication and may be useful as disease biomarkers. We systematically compared small RNAs in 12 human biofluid types using RNA-seq. miRNAs and tRNA-derived RNAs (tDRs) accounted for the majority of mapped reads in all biofluids, but the ratio of miRNA to tDR reads varied from 72 in plasma to 0.004 in bile. miRNA levels were highly correlated across all biofluids but levels of some miRNAs differed markedly between biofluids. tDR populations differed extensively between biofluids. Y RNA fragments were seen in all biofluids and accounted for >10% of reads in blood plasma, serum, and CSF. Reads mapping exclusively to piRNAs were very rare except in seminal plasma. These results demonstrate extensive differences in small RNAs between human biofluids and provide a useful resource for investigating extracellular RNA biology and developing biomarkers.

genomics↗

bioRxiv2017

Accuracy, Reproducibility And Bias Of Next Generation Sequencing For Quantitative Small RNA Profiling: A Multiple Protocol Study Across Multiple Laboratories

Small RNA-seq is increasingly being used for profiling of small RNAs. Quantitative characteristics of long RNA-seq have been extensively described, but small RNA-seq involves fundamentally different methods for library preparation, with distinct protocols and technical variations that have not been fully and systematically studied. We report here the results of a study using common references (synthetic RNA pools of defined composition, as well as plasma-derived RNA) to evaluate the accuracy, reproducibility and bias of small RNA-seq library preparation for five distinct protocols and across nine different laboratories. We observed protocol-specific and sequence-specific bias, which was ameliorated using adapters for ligation with randomized end-nucleotides, and computational correction factors. Despite this technical bias, relative quantification using small RNA-seq was remarkably accurate and reproducible, even across multiple laboratories using different methods. These results provide strong evidence for the feasibility of reproducible cross-laboratory small RNA-seq studies, even those involving analysis of data generated using different protocols.

genomics↗