Biology subjects

Batista, P. J.

Publications and source records attributed to Batista, P. J..

7 recordsLinked to original sources

bioRxiv2026

G quadruplex DNA facilitates a pervasive path to homologous recombination

Homologous recombination (HR) requires efficient homology search and strand invasion, yet how homologous templates are identified within nuclear space remains unclear. Here, we identify G-quadruplex (G4) DNA structures as pervasive effectors of template strand invasion and uncover the G4 helicase DHX36 as a potent suppressor of this process. DHX36 loss stabilizes G4s, enhances HR, and accelerates repair of replication-associated DNA breaks. G4-mediated HR depends on the non-canonical strand invasion factors RAD51AP1, WDR48, and USP1, and requires G4 motifs within the homologous repair template. DHX36 loss partially restores HR and PARP inhibitor resistance in BRCA1-deficient cells, while promoting aberrant recombination and Alternative Lengthening of Telomeres (ALT). Together, our findings establish dynamic G4 regulation as a key determinant of homology search, genome maintenance, and recombination fidelity.

molecular biology↗

bioRxiv2025

A whole genomic CRISPR-Cas9 screen identifies the amino acid transporter SLC43A1 (LAT3) as a major determinant of oxaliplatin sensitivity in colorectal cancer cells

Colorectal cancer (CRC) is the second leading cause of cancer deaths in the United States, with a five-year survival rate of 65%. Oxaliplatin was the first platinum drug shown to improve CRC patient outcomes and is now a common adjuvant therapy for advanced disease, yet 90% of patients develop resistance. Oxaliplatin was developed as a third-generation derivative of cisplatin, but recent evidence points to divergent modes of action. Here, genome-wide CRISPR activation and knockout screens were conducted to identify genetic changes that confer resistance to oxaliplatin in two CRC cell lines with distinct molecular backgrounds (SW620 and RKO). Guide RNAs corresponding to the neutral amino acid transporter SLC43A1 (LAT3) were the most significantly enriched in knockout screens and depleted in activation screens, suggesting a potential role for LAT3 in modulating oxaliplatin resistance. In vitro CRISPR knockout and overexpression of LAT3 in SW620 and RKO cell lines confirm increased resistance or sensitivity to oxaliplatin, respectively. Further analysis demonstrates that increased LAT3 levels corrrelate with increased intracellular levels of oxaliplatin, increased levels of DNA-platinum adducts and DNA damage, demonstrating that enhanced LAT3-mediated uptake of oxaliplatin can exert its expected mechanism of action and induce cytotoxicity. These findings may lead to a better understanding of oxaliplatins mode of action in CRC and can provide new insights into the interplay between essential nutrient uptake and drug transport. STATEMENT OF SIGNIFICANCEOxaliplatin resistance remains a major clinical challenge for CRC treatment. Our study identified a novel role for LAT3 as a modulator of oxaliplatin sensitivity, offering new insights into drug resistance mechanisms.

cancer biology↗

bioRxiv2024

The modification landscape of P. aeruginosa tRNAs

RNA modifications have a substantial impact on tRNA function, with modifications in the anticodon loop contributing to translational fidelity and modifications in the tRNA core impacting structural stability. In bacteria, tRNA modifications are crucial for responding to stress and regulating the expression of virulence factors. Although tRNA modifications are well-characterized in a few model organisms, our knowledge of tRNA modifications in human pathogens, such as Pseudomonas aeruginosa, remains limited. Here we leveraged two orthogonal approaches to build a reference landscape of tRNA modifications in E. coli, which enabled us to identify similar modifications in P. aeruginosa. Our analysis revealed a substantial degree of conservation between the two organisms, while also uncovering potential sites of tRNA modification in P. aeruginosa tRNAs that are not present in E. coli. The mutational signature at one of these sites, position 46 of tRNAGln1(UUG) is dependent on the P. aeruginosa homolog of TapT, the enzyme responsible for the 3-(3-amino-3-carboxypropyl) uridine (acp3U) modification. Identifying which modifications are present on different tRNAs will uncover the pathways impacted by the different tRNA modifying enzymes, some of which play roles in determining virulence and pathogenicity.

microbiology↗

bioRxiv2023

The thiol methyltransferase activity of TMT1A (METTL7A) is conserved across species

Although few resistance mechanisms for histone deacetylase inhibitors (HDACis) have been described, we recently demonstrated that TMT1A (formerly METTL7A) and TMT1B (formerly METTL7B) can mediate resistance to HDACis with a thiol as the zinc-binding group by methylating and inactivating the drug. TMT1A and TMT1B are poorly characterized, and their normal physiological role has yet to be determined. As animal model systems are often used to determine the physiological function of proteins, we investigated whether the ability of these methyltransferases to methylate thiol-based HDACis is conserved across different species. We found that TMT1A was conserved across rats, mice, chickens, and zebrafish, displaying 85.7%, 84.8%, 60.7% and 51.0% amino acid sequence identity, respectively, with human TMT1A. Because TMT1B was not found in the chicken or zebrafish, we focused our studies on the TMT1A homologs. HEK-293 cells were transfected to express mouse, rat, chicken, or zebrafish homologs of TMT1A and all conferred resistance to the thiol-based HDACIs NCH-51, KD-5170 and romidepsin compared to empty vector-transfected cells. Additionally, all homologs blunted the downstream effects of HDACi treatment such as increased p21 expression, increased acetylated histone H3, and cell cycle arrest. Increased levels of dimethylated romidepsin were also found in the culture medium of cells transfected to express any of the TMT1A homologs after a 24 h incubation with romidepsin compared to empty-vector transfected cells. Our results indicate that the ability of TMT1A to methylate molecules is conserved across species. Animal models may therefore be useful in elucidating the role of these enzymes in humans.

pharmacology and toxicology↗

bioRxiv2023

Rewiring of RNA methylation by the oncometabolite fumarate in renal cell carcinoma

Metabolic reprogramming is a hallmark of cancer that facilitates changes in many adaptive biological processes. Mutations in the tricarboxylic acid (TCA) cycle enzyme fumarate hydratase (FH) lead to fumarate accumulation and cause hereditary leiomyomatosis and renal cell cancer (HLRCC). HLRCC is a rare, inherited disease characterized by the development of non-cancerous smooth muscle tumors of the uterus and skin, and an increased risk of a highly metastatic and aggressive form of kidney cancer. Fumarate has been shown to inhibit 2-oxyglutarate-dependent dioxygenases (2OGDDs) involved in the hydroxylation of HIF1, as well as in DNA and histone demethylation. However, the link between fumarate accumulation and changes in RNA post-transcriptional modifications has not been defined. Here, we determine the consequences of fumarate accumulation on the activity of different members of the 2OGDD family targeting RNA modifications. By evaluating multiple RNA modifications in patient-derived HLRCC cell lines, we show that mutation of FH selectively alters the activity of demethylases acting upon N6-methyladenosine (m6A), while the demethylase acting upon N1-methyladenosine (m1A) and 5-formylcytosine (f5C) in mitochondrial RNA are unaffected. The observation that metabolites modulate specific subsets of RNA-modifying enzymes offers new insights into the intersection between metabolism and the epitranscriptome.

molecular biology↗

bioRxiv2022

The methyltransferases METTL7A and METTL7B confer resistance to thiol-based histone deacetylase inhibitors

Histone deacetylase inhibitors (HDACis) are part of a growing class of epigenetic therapies used for the treatment of cancer. While elevated levels of the efflux pump P-gp are associated with in vitro resistance to romidepsin, this mechanism does not translate to the clinic. We developed a romidepsin-resistant cell line with a resistance mechanism independent of P-gp function that acts upstream of the deacetylation process. We found that expression of the methyltransferase METTL7A is necessary for resistance, and that expression of METTL7A in naive cells can drive resistance to thiol-containing HDACis. We demonstrate that METTL7A can methylate romidesin in vitro and that the ability of METTL7A to drive resistance to thiol-containing HDACis can be blocked by the methyltransferase inhibitor DCMB. Our data supports a model whereby exposure of cells to romidepsin selects for upregulation of the methyltransferase METTL7A, which in turn modifies the zinc-binding thiol, inactivating the drug.

cancer biology↗

bioRxiv2019

Functional classification of noncoding RNAs associated with distinct histone modifications by PIRCh-seq

Many long noncoding RNAs (lncRNAs) regulate gene transcription through binding to histone modification complexes. Therefore, a comprehensive study of nuclear RNAs in a histone modification-specific manner is critical to understand their regulatory mechanisms. Here we develop a method named Profiling Interacting RNAs on Chromatin by deep sequencing (PIRCh-seq), in which we profile chromatin-associated transcriptome in 5 different cell types using antibodies recognizing histone H3 and 6 distinct histone modifications associated with active or repressive chromatin states. PIRCh-seq identified chromatin-associated RNAs with substantially less contamination by nascent transcripts, as compared to existing methods. We classified chromatin-enriched lncRNAs into 6 functional groups based on the patterns of their association with specific histone modifications. LncRNAs were enriched with different chromatin modifications in different cell types, suggesting lncRNAs regulation may also be cell type-specific. By integrating profiles of RNA secondary structure and RNA m6A modification, we found that RNA bases which bind to chromatin tend to be more single stranded. We discovered hundreds of allele-specific RNA-chromatin interactions, nominating specific single nucleotide variants that alter RNA association with chromatin. These results provide a unique resource to globally study the functions of chromatin-associated lncRNAs and elucidate the basic mechanisms of chromatin-RNA interaction.

cell biology↗