Biology subjects

Rani, H.

Publications and source records attributed to Rani, H..

8 recordsLinked to original sources

bioRxiv2025

Loss of tumour suppressor p53 rewires enhancer landscape and governs oncogenic progression

Mutations in tumour suppressor p53 confer enhanced metastasis and chemoresistance in colorectal cancer (CRC). Though the genetic events regulating CRC with p53 loss/mutation have been documented, the epigenetic events accompanying the loss of p53 have not been well understood. Epigenome based classification of CRC tumours has identified the active enhancer mark as a distinct marker for progression, however the role of the distal regulatory regions upon p53 loss in CRC remains to be established. This work investigates the influence of p53 loss on enhancer regulation in colorectal cancer cells. Genome wide profiling of active enhancer mark, H3K27ac in p53wt and p53-/- CRC cells reveal an overall gain of this mark around the promoters and intronic regions. These active enhancers show strong association with oncogenes and hallmark MYC and E2F targets suggesting an enhancer mediated regulation of MYC/E2F pathway governed by E2Fs, MAZ and PATZ1. Interestingly, we also observed a gain in oncogenic super enhancers mediated by E2Fs/KLFs accompanying loss of p53. The promoters of histone methyl transferases EZH2 and SuV39H1 (E2F targets) show elevated levels of H3K27ac suggesting a novel epigenetic regulation of CRC around the promoters and distal regulatory regions. Our validation of these findings in p53 deficient colon cancer cohorts shows that the super enhancer associated genes align more to the CMS4 subtype and exhibit lower survivability. The observed cancer stemness and gain of oncogenic super enhancers with p53 loss presents a hitherto unexplored paradigm of enhancer mediated oncogenic progression which may be exploited for devising epigenetic therapy in p53-/- CRC patients. SignificanceColorectal cancers (CRC) lose tumour-suppressor function and gain neomorphic functions with mutation/loss of p53. This work explores the epigenomic modulation of p53 null CRC cells by distal regulatory elements which has not been not clearly understood yet. We report a global increase in the active enhancer mark H3K27ac at active promoter and enhancer regions. We find that the gained enhancers/promoters are regulated by E2Fs/MAZ/PATZ1 which drive cancer stemness while the lost enhancers/promoters are regulated by tumour-suppressive IRFs. The activation of E2Fs correlates with elevated H3K27ac implying positive feedback driving E2F targets such as EZH2 and SuV39H1. The indirect activation of histone methyltransferases by p53 and the gain of oncogenic super-enhancers present a novel epigenetic regulatory paradigm which we also validated in p53 null CRC cohorts. These findings aid the design of epigenetic therapy for p53 deficient colorectal tumours.

genomics↗

bioRxiv2025

Gain of Function p53 mutant R273H confers distinct methylation profiles and consequent partial or full EMT states to colon tumour

p53 is the second most frequently mutated gene in colorectal cancer. While different p53 mutations have been correlated with metastasis, the distinct phenotypes exhibited by site-specific mutations of p53 are not well elucidated. Here, we analyse transcriptomic and methylation data from TCGA-COAD cohort to understand the epigenetic impact of three most prevalent hotspot mutations of p53 (R175H, R273H and R282W). We observed that p53 R273H mutation associates with a partial epithelial-mesenchymal transition (pEMT) state and metastatic progression. In vitro ChIP-seq experiments conducted on p53R27H harbouring HT29 cells revealed an enrichment of mutant p53 R273H at pEMT or mesenchymal gene sets. Further, simulations from a gene regulatory network incorporating the interactions of p53R273H with EMT regulators explain how this mutation shapes the phenotypic landscape accessible to cancer cells. Finally, single-cell transcriptomic analysis of colorectal tumours reveals R273H-linked enrichment of partial and mesenchymal EMT phenotypes across tumour subpopulations in CRC. Overall, we identified distinct epigenetic regulation regulating partial EMT and consequent aggressive behaviour triggered by p53R273H. These findings can help devise effective therapeutic strategies for p53 mutant specific colon tumours. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=177 SRC="FIGDIR/small/662954v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@7d424dorg.highwire.dtl.DTLVardef@18bfcaborg.highwire.dtl.DTLVardef@17950c8org.highwire.dtl.DTLVardef@1a698f9_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical Abstract CaptionC_FLOATNO Mutation in Tp53 at R273H associates with DNA hypermethylation and elevated pEMT and metastatic signatures as compared to R175 and R282W mutations in colon tumours suggesting of devising novel therapeutic intervention strategies based on p53 mutation profiles. C_FIG

bioinformatics↗

bioRxiv2025

Proteomics-based models of gene expression and cellular control of cotton fiber development

The shapes and material properties of cotton seed coat trichoblasts are the basis of a multibillion-dollar natural fiber industry. As such, these highly specialized cells are low-hanging fruit for intentional trait engineering. However, broad successes will require more mechanistic knowledge about their systems-level cellular controls. This time-series study integrates daily measurements of purified fiber transcriptomes and proteomes with multiscale fiber phenotyping datasets that span the same developmental interval. Abundance profiles of the subcellular proteomes are the foundation of the analyses. This resource article provides direct information concerning which homoeologs operate and informative depictions of how compartmentalized cellular systems change during developmental transitions. Prediction accuracy was partially validated by analysis of the protein expression group 11, which contained multiple known secondary cell wall cellulose synthases and dozens of unknown proteins and an averaged profile that was strongly correlated with a sharp state transition in cellulose microfibril alignment and increased cellulose content. The dataset as a whole can serve as a hypothesis-generating machine to guide future experiments that relate to cell shape and growth rate control, reversible tissue formation, and cell wall remodeling. Integration of mRNA and protein abundance revealed widespread evidence for post-transcriptional control. In addition, there were hundreds of transcriptionally controlled genes with differing timepoints of transition. This latter gene set can be used to more reliably analyze transcriptional control networks and to generate collections of gene expression drivers for cotton fiber research. The protein and transcript data are organized into user-friendly tables and a web interface that can be searched using any plant ortholog of interest based on developmental time, abundance, annotations, or phenotypic association.

plant biology↗

bioRxiv2025

Development and quality assessment of low-cost benchtop malting protocol for laboratory-scale malt quality evaluation

High-quality malt is influenced by three primary factors: barley genotype, environmental conditions, and malting process. To effectively evaluate malting barley breeding material and assess how environmental changes influence malt quality, it is essential to have laboratory- scale malting methods that can produce malt approximating that produced by commercial malting operations. However, existing laboratory-scale malting procedures often demand large quantities of grain, rely on specialized equipment, and are costly. To overcome these challenges, we developed a small sample-scale benchtop malting method utilizing standard laboratory equipment and components available at hardware stores. We validated the method by conducting standard malt quality tests including diastatic power, -amylase activity, total malt protein, and wort composition (soluble protein, wort soluble/total malt protein, {beta}-glucan, free amino nitrogen, and malt extract). Our findings indicate that the benchtop malting method yields quality metrics comparable to those obtained from established small-scale and full-scale malting protocols. Furthermore, a key innovation of this system is the use of separate Erlenmeyer flasks for malting each sample. Unlike conventional shared malting systems, this design enables precise measurement and comparison of treatment effects across samples malted simultaneously. This reliable, low-cost, and efficient method provides a valuable tool for screening malt quality traits in breeding lines with limited sample sizes and for testing malting regimes aimed at improving malt quality and efficiency. Additionally, it offers an accessible solution for producing high-quality, research-scale malt in laboratories without dedicated quality assurance facilities.

biochemistry↗

bioRxiv2024

Developmental variability in cotton fiber cell wall properties linked to important agronomic traits

The economic value of cotton is based on its long, thin, strong, and twisted trichoblasts that emerge from the ovule epidermis. The mature dried fiber cell reflects the outcome of a rapid tapering of the nascent trichoblast, weeks of polarized diffuse growth, followed by a transition to persistent secondary cell wall synthesis. Highly conserved and dynamic microtubule and cellulose microfibril-based anisotropic growth control modules are central to all of these phases. In this paper, we developed novel quantitative phenotyping and computational modeling pipelines to analyze fiber growth behaviors at a daily resolution. We uncovered unexpected variability in growth rate, cell wall properties, and cell geometry across a critical window of fiber development. Finite element computational modeling of fiber growth was used to analyze the instability of cell diameter control and predict how spatial gradients of fiber and matrix material properties can interact to dictate the patterns of shape change. As an initial step toward gaining insight into the molecular orchestration of cellulose biosynthesis, expression profiles of a broad set of relevant genes were quantified across the same developmental timeline and correlated with fiber phenotypes. This analysis identified specific candidate genes that may serve as targets for fiber quality improvement.

plant biology↗

bioRxiv2024

A cell fractionation and quantitative proteomics pipeline to enable functional analyses of cotton fiber development

Cotton fibers are aerial trichoblasts that employ a highly polarized diffuse growth mechanism to emerge from the developing ovule epidermis. After executing a complicated morphogenetic program, the cells reach lengths over 2 cm and serve as the foundation of a multi-billion-dollar textile industry. Important traits such as fiber diameter, length, and strength are defined by the growth patterns and cell wall properties of individual cells. At present, the ability to engineer fiber traits is limited by our lack of understanding regarding the primary controls governing the rate, duration, and patterns of cell growth. To gain insights into the compartmentalized functions of proteins in cotton fiber cells, we developed a label-free liquid chromatography mass spectrometry method for systems level analyses of fiber proteome. Purified fibers from a single locule were used to fractionate the fiber proteome into apoplast (APOT), membrane-associated (p200), and crude cytosolic (s200) fractions. Subsequently, proteins were identified, and their localizations and potential functions were analyzed using combinations of size exclusion chromatography, statistical and bioinformatic analyses. This method had good coverage of the p200 and apoplast fractions, the latter of which was dominated by proteins associated with particulate membrane-enclosed compartments. The apoplastic proteome was diverse, the proteins were not degraded, and some displayed distinct multimerization states compared to their cytosolic pool. This quantitative proteomic pipeline can be used to improve coverage and functional analyses of the cotton fiber proteome as a function of developmental time or differing genotypes.

plant biology↗

bioRxiv2024

A high-resolution model of gene expression during Gossypium hirsutum (cotton) fiber development

Cotton fiber development relies on complex and intricate biological processes to transform newly differentiated fiber initials into the mature, extravagantly elongated cellulosic cells that are the foundation of this economically important cash crop. Here we extend previous research into cotton fiber development by employing controlled conditions to minimize variability and utilizing time-series sampling and analyses to capture daily transcriptomic changes from early elongation through the early stages of secondary wall synthesis (6 to 24 days post anthesis; DPA). A majority of genes are expressed in fiber, largely partitioned into two major coexpression modules that represent genes whose expression generally increases or decreases during development. Differential gene expression reveals a massive transcriptomic shift between 16 and 17 DPA, corresponding to the onset of the transition phase that leads to secondary wall synthesis. Subtle gene expression changes are captured by the daily sampling, which are discussed in the context of fiber development. Coexpression and gene regulatory networks are constructed and associated with phenotypic aspects of fiber development, including turgor and cellulose production. Key genes are considered in the broader context of plant secondary wall synthesis, noting their known and putative roles in cotton fiber development. The analyses presented here highlight the importance of fine-scale temporal sampling on understanding developmental processes and offer insight into genes and regulatory networks that may be important in conferring the unique fiber phenotype.

cell biology↗

bioRxiv2024

Integrative LC-MS and GC-MS Metabolic Profiling Unveils Dynamic Changes during Barley Malting

Malting, a crucial process for beer production, involves complex biochemical transformations affecting sensory attributes and product quality. Despite extensive research on storage carbohydrates and proteins involved in malting, a detailed understanding of metabolic alterations during this process remains elusive, limiting our ability to assess and enhance malt quality. Our study employed untargeted GC-MS and LC-MS metabolite profiling to elucidate these changes across six malting stages: dry seed, post-steeping (DOG0), germination (DOG1, DOG3, DOG5), and kilning. We identified a total of 4980 known metabolites, with approximately 82% exhibiting significant changes. Statistical analysis revealed stage-dependent metabolic shifts, with most significant shifts occurring from DOG1 to DOG3 and during kilning. Dynamic changes in various chemical classes and metabolic pathways provide insights into processes critical for malt quality and beer production. Additionally, metabolites associated with antimicrobial properties and stress responses were identified, underscoring the interplay between barley and microbial metabolic processes during malting. HighlightsO_LIGC-MS and LC-MS profiling were performed to track metabolic changes during malting. C_LIO_LIIdentified 4980 known compounds belonging to 346 chemical classes during malting. C_LIO_LIMany microbial metabolites demonstrated increased abundance in finished malt. C_LIO_LIThe most significant metabolic shifts occurred during early germination and kilning. C_LI

biochemistry↗