BORIS/CTCFL-mediated chromatin accessibility alterations promote a pro-invasive transcriptional signature in melanoma cells.

Melanoma is the deadliest form of skin cancer, due to its tendency to metastasize early. Brother of regulator of imprinted sites (BORIS), also known as CCCTC binding factor-like (CTCFL), is a transcription regulator that becomes ectopically expressed in melanoma. We recently showed that BORIS contributes to melanoma phenotype switching by altering the gene expression program of melanoma cells from an intermediate melanocytic state toward a more mesenchymal-like state. However, the mechanism underlying this transcriptional switch remains unclear. Here, ATAC-seq was used to study BORIS-mediated chromatin accessibility alterations in melanoma cells harboring an intermediate melanocytic state. The gene set that gained promoter accessibility, following ectopic BORIS expression, showed enrichment for biological processes associated with melanoma invasion, while promoters of genes associated with proliferation showed reduced accessibility. Integration of ATAC-seq and RNA-seq data demonstrated that increased chromatin accessibility was associated with transcriptional upregulation of genes involved in tumor progression processes, and the aberrant activation of oncogenic transcription factors, while reduced chromatin accessibility and downregulated genes were associated with repressed activity of tumor suppressors and proliferation factors. Together, these findings indicate that BORIS mediates transcriptional reprogramming in melanoma cells by altering chromatin accessibility and gene expression, shifting the cellular transcription landscape of melanoma cells toward a mesenchymal-like genetic signature.

BORIS/CTCFL promotes a switch from a proliferative towards an invasive phenotype in melanoma cells.

Melanoma is among the most aggressive cancers due to its tendency to metastasize early. Phenotype switching between a proliferative and an invasive state has been suggested as a critical process for metastasis, though the mechanisms that regulate state transitions are complex and remain poorly understood. Brother of Regulator of Imprinted Sites (BORIS), also known as CCCTC binding factor-Like (CTCFL), is a transcriptional modulator that becomes aberrantly expressed in melanoma. Yet, the role of BORIS in melanoma remains elusive. Here, we show that BORIS is involved in melanoma phenotype switching. Genetic modification of BORIS expression in melanoma cells combined with whole-transcriptome analysis indicated that BORIS expression contributes to an invasion-associated transcriptome. In line with these findings, inducible BORIS overexpression in melanoma cells reduced proliferation and increased migration and invasion, demonstrating that the transcriptional switch is accompanied by a phenotypic switch. Mechanistically, we reveal that BORIS binds near the promoter of transforming growth factor-beta 1 (TFGB1), a well-recognized factor involved in the transition towards an invasive state, which coincided with increased expression of TGFB1. Overall, our study indicates a pro-invasive role for BORIS in melanoma via transcriptional reprogramming.

Comparison between next-generation and Sanger-based sequencing for the detection of transmitted drug-resistance mutations among recently infected HIV-1 patients in Israel, 2000-2014.

INTRODUCTION: Transmitted drug-resistance mutations (TDRM) may hamper successful anti-HIV-1 therapy and impact future control of the HIV-1 epidemic. Recently infected, therapy-naïve individuals are best suited for surveillance of such TDRM. In this study, TDRM, detected by next-generation sequencing (NGS) were compared to those identified by Sanger-based population sequencing (SBS) in recently infected HIV-1 patients. METHODS: Historical samples from 80 recently infected HIV-1 patients, diagnosed between 2000 and 2014, were analysed by MiSeq (NGS) and ABI (SBS). DeepChek-HIV (ABL) was used for interpretation of the results. RESULTS: Most patients were males (80%); Men who have sex with men (MSM) was the major transmission group (58.8%). Overall, TDRM were detected in 31.3% of patients by NGS and 8.8% by SBS, with SBS TDRM restricted to persons infected with subtype B. All SBS-detected TDRM were identified by NGS. The prevalence of TDRM impacting protease inhibitors (PI), nucleoside reverse transcriptase inhibitors (NRTI) and non-nucleoside reverse transcriptase inhibitors (NNRTI) was 11.3, 26.2 7.5%, respectively, in NGS analyses and 0, 3.8 and 5%, respectively, in SBS analyses. More patients with NGS and SBS TDRM were identified in 2008-2014 (37.2% or 13.9%, respectively) compared to 2000-2007 (24.3% or 2.7%, respectively), and a significantly greater number of these patients had multiple NGS TDRM. The most abundant, albeit, minor-frequency RT TDRM, were the K65R and D67N, while K103N, M184V and T215S were high-frequency mutations. Minor TDRM did not become a major variant in later samples and did not hinder successful treatment. CONCLUSIONS: NGS can replace SBS for mutation detection and allows for the detection of low-frequency TDRM not identified by SBS. Although rates of TDRM in Israel continued to increase from 2000 to 2014, minor TDRM did not become major species. The need for ongoing surveillance of low-frequency TDRM should be revisited in a larger study.

HCV genotype-1 subtypes and resistance-associated substitutions in drug-naive and in direct-acting antiviral treatment failure patients.

BACKGROUND: Direct-acting antiviral (DAA) treatment regimens and response rates of patients with HCV genotype-1 (GT1) are currently considered subtype-dependent. Identification of clinically relevant resistance-associated substitutions (RASs) in the NS3 and NS5A proteins at baseline and in DAA failures, may also impact clinical decisions. METHODS: In a multicentre cohort study (n=308), NS3 or NS5B sequencing (n=248) was used to discriminate between GT1 subtypes. The correlation between baseline NS3 and NS5A RASs on the 12-week sustained virological response (SVR12) rates of 160 of the patients treated with second-generation DAAs was also assessed. Post-treatment resistance analysis was performed on samples from 58 patients exhibiting DAA virological failure. RESULTS: GT1a, GT1b and GT1d subtypes were identified in 23.0%, 75.4% and 1.2% of tested samples. GT1b was most prevalent (97.7%, 128/131) among patients born in the former Soviet Union. The Q80K NS3 RAS was identified in 17.5% (10/57) of the GT1a carriers, most of whom were Israeli-born. NS3 and NS5A baseline RASs showed a negligible correlation with SVR12 rates. Treatment-emergent RASs were observed among 8.9% (4/45) and 76.9% (10/13) of first- and second-generation DAA failures, respectively, with D168V/E (NS3), Y93H and L31M (NS5A) being the most prevalent mutations. CONCLUSIONS: NS3 sequencing analysis can successfully discriminate between GT1 subtypes and identify NS3 amino acid substitutions. While pre-treatment NS3 and NS5A RASs marginally affect second-generation DAA SVR12 rates, post-treatment resistance analysis should be considered prior to re-therapy.