@article {123130, title = {Active enhancers strengthen insulation by RNA-mediated CTCF binding at chromatin domain boundaries.}, journal = {Genome Res}, year = {2023}, month = {2023 Jan 17}, abstract = {

Vertebrate genomes are partitioned into chromatin domains or topologically associating domains (TADs), which are typically bound by head-to-head pairs of CTCF binding sites. Transcription at domain boundaries correlates with better insulation; however, it is not known whether the boundary transcripts themselves contribute to boundary function. Here we characterize boundary-associated RNAs genome-wide, focusing on the disease-relevant and TAD. Using CTCF site deletions and boundary-associated RNA knockdowns, we observe that boundary-associated RNAs facilitate recruitment and clustering of CTCF at TAD borders. The resulting CTCF enrichment enhances TAD insulation, enhancer-promoter interactions, and TAD gene expression. Importantly, knockdown of boundary-associated RNAs results in loss of boundary insulation function. Using enhancer deletions and CRISPRi of promoters, we show that active TAD enhancers, but not promoters, induce boundary-associated RNA transcription, thus defining a novel class of regulatory enhancer RNAs.

}, issn = {1549-5469}, doi = {10.1101/gr.276643.122}, author = {Islam, Zubairul and Saravanan, Bharath and Walavalkar, Kaivalya and Farooq, Umer and Singh, Anurag Kumar and Sabarinathan, Radhakrishnan and Thakur, Jitendra and Pandit, Awadhesh and Henikoff, Steven and Notani, Dimple} } @article {136529, title = {Cis-regulatory effect of HPV integration is constrained by host chromatin architecture in cervical cancers.}, journal = {Mol Oncol}, year = {2023}, month = {2023 Nov 27}, abstract = {

HPV infections are the primary drivers of cervical cancers, and often HPV DNA gets integrated into the host genome. Although the oncogenic impact of HPV encoded genes is relatively well known, the cis-regulatory effect of integrated HPV DNA on host chromatin structure and gene regulation remains less understood. We investigated genome-wide patterns of HPV integrations and associated host gene expression changes in the context of host chromatin states and TADs. HPV integrations were significantly enriched in active chromatin regions and depleted in inactive ones. Interestingly, regardless of chromatin state, genomic regions flanking HPV integrations showed transcriptional upregulation. Nevertheless, upregulation (both local and long-range) was mostly confined to TADs with integration, but not affecting adjacent TADs. Few TADs showed recurrent integrations associated with overexpression of oncogenes within them (e.g. MYC, PVT1, TP63 and ERBB2) regardless of proximity. Hi-C and 4C-seq analyses in cervical cancer cell line (HeLa) demonstrated chromatin looping interactions between integrated HPV and MYC/PVT1 regions (~500kb apart), leading to allele-specific overexpression. Based on these, we propose HPV integrations can trigger multimodal oncogenic activation to promote cancer progression.

}, issn = {1878-0261}, doi = {10.1002/1878-0261.13559}, author = {Singh, Anurag Kumar and Walavalkar, Kaivalya and Tavernari, Daniele and Ciriello, Giovanni and Notani, Dimple and Sabarinathan, Radhakrishnan} } @article {129160, title = {Editing the core region in HPFH deletions alters fetal and adult globin expression for treatment of β-hemoglobinopathies.}, journal = {Mol Ther Nucleic Acids}, volume = {32}, year = {2023}, month = {2023 Jun 13}, pages = {671-688}, abstract = {

Reactivation of fetal hemoglobin (HbF) is a commonly adapted strategy to ameliorate β-hemoglobinopathies. However, the continued production of defective adult hemoglobin (HbA) limits HbF tetramer production affecting the therapeutic benefits. Here, we evaluated deletional hereditary persistence of fetal hemoglobin (HPFH) mutations and identified an 11-kb sequence, encompassing putative repressor region (PRR) to β-globin exon-1 (βE1), as the core deletion that ablates HbA and exhibits superior HbF production compared with HPFH or other well-established targets. PRR-βE1-edited hematopoietic stem and progenitor cells (HSPCs) retained their genome integrity and their engraftment potential to repopulate for long-term hematopoiesis in immunocompromised mice producing HbF positive cells . Furthermore, PRR-βE1 gene editing is feasible without HSPC culture. Importantly, the editing induced therapeutically significant levels of HbF to reverse the phenotypes of both sickle cell disease and β-thalassemia major. These findings imply that PRR-βE1 gene editing of patient HSPCs could lead to improved therapeutic outcomes for β-hemoglobinopathy gene therapy.

}, issn = {2162-2531}, doi = {10.1016/j.omtn.2023.04.024}, author = {Venkatesan, Vigneshwaran and Christopher, Abisha Crystal and Rhiel, Manuel and Azhagiri, Manoj Kumar K and Babu, Prathibha and Walavalkar, Kaivalya and Saravanan, Bharath and Andrieux, Geoffroy and Rangaraj, Sumathi and Srinivasan, Saranya and Karuppusamy, Karthik V and Jacob, Annlin and Bagchi, Abhirup and Pai, Aswin Anand and Nakamura, Yukio and Kurita, Ryo and Balasubramanian, Poonkuzhali and Pai, Rekha and Marepally, Srujan Kumar and Mohankumar, Kumarasamypet Murugesan and Velayudhan, Shaji R and Boerries, Melanie and Notani, Dimple and Cathomen, Toni and Srivastava, Alok and Thangavel, Saravanabhavan} } @article {113537, title = {Recommendations for empowering early career researchers to improve research culture and practice.}, journal = {PLoS Biol}, volume = {20}, year = {2022}, month = {2022 Jul 07}, pages = {e3001680}, abstract = {

Early career researchers (ECRs) are important stakeholders leading efforts to catalyze systemic change in research culture and practice. Here, we summarize the outputs from a virtual unconventional conference (unconference), which brought together 54 invited experts from 20 countries with extensive experience in ECR initiatives designed to improve the culture and practice of science. Together, we drafted 2 sets of recommendations for (1) ECRs directly involved in initiatives or activities to change research culture and practice; and (2) stakeholders who wish to support ECRs in these efforts. Importantly, these points apply to ECRs working to promote change on a systemic level, not only those improving aspects of their own work. In both sets of recommendations, we underline the importance of incentivizing and providing time and resources for systems-level science improvement activities, including ECRs in organizational decision-making processes, and working to dismantle structural barriers to participation for marginalized groups. We further highlight obstacles that ECRs face when working to promote reform, as well as proposed solutions and examples of current best practices. The abstract and recommendations for stakeholders are available in Dutch, German, Greek (abstract only), Italian, Japanese, Polish, Portuguese, Spanish, and Serbian.

}, issn = {1545-7885}, doi = {10.1371/journal.pbio.3001680}, author = {Kent, Brianne A and Holman, Constance and Amoako, Emmanuella and Antonietti, Alberto and Azam, James M and Ballhausen, Hanne and Bediako, Yaw and Belasen, Anat M and Carneiro, Clarissa F D and Chen, Yen-Chung and Compeer, Ewoud B and Connor, Chelsea A C and Cr{\"u}well, Sophia and Debat, Humberto and Dorris, Emma and Ebrahimi, Hedyeh and Erlich, Jeffrey C and Fern{\'a}ndez-Chiappe, Florencia and Fischer, Felix and Gazda, Ma{\l}gorzata Anna and Glatz, Toivo and Grabitz, Peter and Heise, Verena and Kent, David G and Lo, Hung and McDowell, Gary and Mehta, Devang and Neumann, Wolf-Julian and Neves, Kleber and Patterson, Mark and Penfold, Naomi C and Piper, Sophie K and Puebla, Iratxe and Quashie, Peter K and Quezada, Carolina Paz and Riley, Julia L and Rohmann, Jessica L and Saladi, Shyam and Schwessinger, Benjamin and Siegerink, Bob and Stehlik, Paulina and Tzilivaki, Alexandra and Umbers, Kate D L and Varma, Aalok and Walavalkar, Kaivalya and de Winde, Charlotte M and Zaza, Cecilia and Weissgerber, Tracey L} } @article {91958, title = {Creating clear and informative image-based figures for scientific publications.}, journal = {PLoS Biol}, volume = {19}, year = {2021}, month = {2021 Mar 31}, pages = {e3001161}, abstract = {

Scientists routinely use images to display data. Readers often examine figures first; therefore, it is important that figures are accessible to a broad audience. Many resources discuss fraudulent image manipulation and technical specifications for image acquisition; however, data on the legibility and interpretability of images are scarce. We systematically examined these factors in non-blot images published in the top 15 journals in 3 fields; plant sciences, cell biology, and physiology (n = 580 papers). Common problems included missing scale bars, misplaced or poorly marked insets, images or labels that were not accessible to colorblind readers, and insufficient explanations of colors, labels, annotations, or the species and tissue or object depicted in the image. Papers that met all good practice criteria examined for all image-based figures were uncommon (physiology 16\%, cell biology 12\%, plant sciences 2\%). We present detailed descriptions and visual examples to help scientists avoid common pitfalls when publishing images. Our recommendations address image magnification, scale information, insets, annotation, and color and may encourage discussion about quality standards for bioimage publishing.

}, issn = {1545-7885}, doi = {10.1371/journal.pbio.3001161}, author = {Jambor, Helena and Antonietti, Alberto and Alicea, Bradly and Audisio, Tracy L and Auer, Susann and Bhardwaj, Vivek and Burgess, Steven J and Ferling, Iuliia and Gazda, Ma{\l}gorzata Anna and Hoeppner, Luke H and Ilangovan, Vinodh and Lo, Hung and Olson, Mischa and Mohamed, Salem Yousef and Sarabipour, Sarvenaz and Varma, Aalok and Walavalkar, Kaivalya and Wissink, Erin M and Weissgerber, Tracey L} } @article {91470, title = {An interdependent network of functional enhancers regulates transcription and EZH2 loading at the INK4a/ARF locus.}, journal = {Cell Rep}, volume = {34}, year = {2021}, month = {2021 Mar 23}, pages = {108898}, abstract = {

The INK4a/ARF locus encodes important cell-cycle regulators p14, p15, and p16. The neighboring gene desert to this locus is the most reproducible GWAS hotspot that harbors one of the densest enhancer clusters in the genome. However, how multiple enhancers that overlap with GWAS variants regulate the INK4a/ARF locus is unknown, which is an important step in linking genetic variation with associated diseases. Here, we show that INK4a/ARF promoters interact with a subset of enhancers in the cluster, independent of their H3K27ac and eRNA levels. Interacting enhancers transcriptionally control each other and INK4a/ARF promoters over long distances as an interdependent single unit. The deletion of even a single interacting enhancer results in an unexpected collapse of the entire enhancer cluster and leads to EZH2 enrichment on promoters in an ANRIL-independent manner. Dysregulated genes genome-wide mimic 9p21-associated diseases under these scenarios. Our results highlight intricate dependencies of promoter-interacting enhancers on each other.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2021.108898}, author = {Farooq, Umer and Saravanan, Bharath and Islam, Zubairul and Walavalkar, Kaivalya and Singh, Anurag Kumar and Jayani, Ranveer Singh and Meel, Sweety and Swaminathan, Sudha and Notani, Dimple} } @article {78108, title = {Beyond the coding genome: non-coding mutations and cancer.}, journal = {Front Biosci (Landmark Ed)}, volume = {25}, year = {2020}, month = {2020 Jun 01}, pages = {1828-1838}, abstract = {

Latest advancements in genomics involving individuals from different races and geographical locations has led to the identification of thousands of common as well as rare genetic variants and copy number variations (CNVs). These studies have surprisingly revealed that the majority of genetic variation is not present within the coding region but rather in the non-coding region of the genome, which is also termed as "Medical Genome". This short review describes how mutations/variations within; regulatory sequences, architectural proteins and transcriptional regulators give rise to the aberrant gene expression profiles that drives cellular transformations and malignancies.

}, issn = {1093-4715}, author = {Walavalkar, Kaivalya and Notani, Dimple} } @article {71098, title = {Ligand dependent gene regulation by transient ERα clustered enhancers.}, journal = {PLoS Genet}, volume = {16}, year = {2020}, month = {2020 Jan 06}, pages = {e1008516}, abstract = {

Unliganded Estrogen receptor alpha (ERα) has been implicated in ligand-dependent gene regulation. Upon ligand exposure, ERα binds to several EREs relatively proximal to the pre-marked, or persistent, ERα-bound sites and affects transient but robust gene expression. However, the underlying mechanisms are not fully understood. Here we demonstrate that upon ligand stimulation, persistent sites interact extensively, via chromatin looping, with the proximal transiently ERα-bound sites, forming Ligand Dependent ERα Enhancer Cluster in 3D (LDEC). The E2-target genes are regulated by these clustered enhancers but not by the H3K27Ac super-enhancers. Further, CRISPR-based deletion of TFF1 persistent site disrupts the formation of its LDEC resulting in the loss of E2-dependent expression of TFF1 and its neighboring genes within the same TAD. The LDEC overlap with nuclear ERα condensates that coalesce in a ligand and persistent site dependent manner. Furthermore, formation of clustered enhancers, as well as condensates, coincide with the active phase of signaling and their later disappearance results in the loss of gene expression even though persistent sites remain bound by ERα. Our results establish, at TFF1 and NRIP1 locus, a direct link between ERα condensates, ERα enhancer clusters, and transient, but robust, gene expression in a ligand-dependent fashion.

}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1008516}, author = {Saravanan, Bharath and Soota, Deepanshu and Islam, Zubairul and Majumdar, Sudeshna and Mann, Rajat and Meel, Sweety and Farooq, Umer and Walavalkar, Kaivalya and Gayen, Srimonta and Singh, Anurag Kumar and Hannenhalli, Sridhar and Notani, Dimple} } @article {79800, title = {A rare variant of African ancestry activates 8q24 lncRNA hub by modulating cancer associated enhancer.}, journal = {Nat Commun}, volume = {11}, year = {2020}, month = {2020 Jul 17}, pages = {3598}, abstract = {

Genetic variation at the 8q24 locus is linked with the greater susceptibility to prostate cancer in men of African ancestry. One such African ancestry specific rare variant, rs72725854 (A\>G/T) (~6\% allele frequency) has been associated with a ~2-fold increase in prostate cancer risk. However, the functional relevance of this variant is unknown. Here we show that the variant rs72725854 is present in a prostate cancer-specific enhancer at 8q24 locus. Chromatin-conformation capture and dCas9 mediated enhancer blocking establish a direct regulatory link between this enhancer and lncRNAs PCAT1, PRNCR1 and PVT1. The risk allele ({\textquoteright}T{\textquoteright}) is associated with higher expression of PCAT1, PVT1 and c-myc in prostate tumors. Further, enhancer with the risk allele gains response to androgen stimulation by recruiting the transcription factor SPDEF whereas, non-risk alleles remain non-responsive. Elevated expression of these lncRNAs and c-myc in risk allele carriers may explain their greater susceptibility to prostate cancer.

}, issn = {2041-1723}, doi = {10.1038/s41467-020-17325-y}, author = {Walavalkar, Kaivalya and Saravanan, Bharath and Singh, Anurag Kumar and Jayani, Ranveer Singh and Nair, Ashwin and Farooq, Umer and Islam, Zubairul and Soota, Deepanshu and Mann, Rajat and Shivaprasad, Padubidri V and Freedman, Matthew L and Sabarinathan, Radhakrishnan and Haiman, Christopher A and Notani, Dimple} }