Biology subjects

Chauhan, V. P.

Publications and source records attributed to Chauhan, V. P..

3 recordsLinked to original sources

bioRxiv2024

Mutant p53 Exploits Enhancers to Elevate Immunosuppressive Chemokine Expression and Impair Immune Checkpoint Inhibitors in Pancreatic Cancer

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer without effective treatments. It is characterized by activating KRAS mutations and p53 alterations. However, how these mutations dysregulate cancer-cell-intrinsic gene programs to influence the immune landscape of the tumor microenvironment (TME) remains poorly understood. Here, we show that p53R172H establishes an immunosuppressive TME, diminishes the efficacy of immune checkpoint inhibitors (ICIs), and enhances tumor growth. Our findings reveal that the upregulation of the immunosuppressive chemokine Cxcl1 mediates these pro-tumorigenic functions of p53R172H. Mechanistically, we show that p53R172H associates with the distal enhancers of the Cxcl1 gene, increasing enhancer activity and Cxcl1 expression. p53R172H occupies these enhancers in an NF-{kappa}B-pathway-dependent manner, suggesting NF-{kappa}Bs role in recruiting p53R172H to the Cxcl1 enhancers. Our work uncovers how a common mutation in a tumor-suppressor transcription factor appropriates enhancers, stimulating chemokine expression and establishing an immunosuppressive TME that diminishes ICI efficacy in PDAC. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/609802v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@b2354dorg.highwire.dtl.DTLVardef@1c96300org.highwire.dtl.DTLVardef@829f26org.highwire.dtl.DTLVardef@19e7d2e_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

Engineered prime editors with minimal genomic errors

Prime editors make programmed genome modifications by writing new sequences into extensions of nicked DNA 3 ends. These edited 3 new strands must displace competing 5 strands to install edits, yet a bias toward retaining the competing 5 strands hinders efficiency and can cause indel errors. Using rational design of the constituent Cas9-nickase to reposition prime editor nicks, we discovered that competing 5 strands are destabilized to favor the edited 3 new strands. We exploit this mechanism to engineer efficient prime editors with strikingly low indel errors. Combining this error-suppressing strategy with the latest efficiency-boosting architecture, we design a next- generation prime editor (vPE). Compared with previous editors, vPE features comparable efficiency yet up to 60-fold lower indel errors, enabling edit:indel ratios as high as 465:1. One Sentence SummaryPrime editors designed with repositioned DNA breaks nearly eliminate undesired genome editing errors

bioengineering↗

bioRxiv2022

Altered DNA repair pathway engagement by engineered CRISPR-Cas9 nucleases

CRISPR-Cas9 introduces targeted DNA breaks that engage competing DNA repair pathways, producing a spectrum of imprecise insertion/deletion mutations (indels) and precise templated mutations (precise edits). The relative frequencies of these pathways are thought to primarily depend on genomic sequence and cell state contexts, limiting control over mutational outcomes. Here we report that engineered Cas9 nucleases that create different DNA break structures engage competing repair pathways at dramatically altered frequencies. We accordingly designed a Cas9 variant (vCas9) that produces breaks which suppress otherwise dominant nonhomologous end-joining (NHEJ) repair. Instead, breaks created by vCas9 are predominantly repaired by pathways utilizing homologous sequences, specifically microhomology-mediated end-joining (MMEJ) and homology-directed repair (HDR). Consequently, vCas9 enables efficient precise editing through HDR or MMEJ while suppressing indels caused by NHEJ in dividing and non-dividing cells. These findings establish a new paradigm of targeted nucleases custom-designed for specific mutational applications. TeaserCRISPR-Cas9 can be designed to make otherwise infrequent precise editing pathways dominant in dividing and non-dividing cells

bioengineering↗