Biology subjects

Subramanian, H.

Publications and source records attributed to Subramanian, H..

11 recordsLinked to original sources

bioRxiv2026

Sequence-Dependent DNA Base Selection Fidelity: A Kinetics-based Model and its validation

DNA replication achieves error rates as low as 10-9-10-11 per base pair through the combined contribution of base selection, exonuclease proofreading, and mismatch repair. Among those processes, base selection determines the initial level of accuracy and is strongly modulated by local sequence context. Existing models address this dependence either by fitting individual rate constants for each sequence context or by invoking the global template properties, neither of which derives sequence dependence from the underlying thermodynamics and kinetics of base pair formation. Here we present a mechanism for sequence-dependent base selection fidelity, built from two physical properties: nearest-neighbor stacking thermodynamics and directional kinetic asymmetry. The model fits the experimentally observed mutation spectra from three mismatch repair-deficient organisms well (r=0.74, 0.70, and 0.63), and predicts that base-selection accuracy varies non-monotonically with temperature in a sequence-dependent manner. Our model, therefore, provides a framework that connects sequence-dependent thermodynamic and kinetic effects during nucleotide incorporation to experimentally observed mutation rates.

biochemistry↗

bioRxiv2026

Directional purine pyrimidine asymmetry in CRISPR repeats facilitates spacer acquisition

Spacer acquisition is the primary and essential step of CRISPR-Cas adaptive immunity in most prokaryotes and occurs preferentially at the leader-repeat junction of the CRISPR array. Despite the conservation of the adaptation machinery, spacer acquisition efficiencies and site-specificity vary markedly across CRISPR-Cas systems, suggesting that the local sequence architecture of the repeat and leader-repeat junction may contribute to the variation in the adaptation efficiency. To investigate this possibility, we systematically analyzed the direct repeat sequences and the terminal base pairs at the 3' end of leader regions adjacent to the integration site. We identify a conserved asymmetric purine-pyrimidine (RY) distribution within direct repeats, characterized by a 5' pyrimidine-rich half and a 3' purine-rich half, together with purine enrichment at the 3' end of leader sequences. Building on these observations, we develop a mechanistic model for spacer acquisition by demonstrating the importance of the sequence motif of the first direct repeat and the leader-repeat junction, using the sequence-dependent asymmetric cooperativity model that captures DNA unzipping kinetics. According to our model, sequences with such directional RY-asymmetric nucleotide distribution as direct repeats have high unzipping propensity and thus can promote efficient spacer integration. Consistent with this mechanism, experimental data from previous studies show that repeats with stronger asymmetry and appropriately positioned pyrimidine-to-purine transition sites are associated with larger spacer counts within CRISPR arrays. These findings reveal a conserved architectural feature of CRISPR repeats and provide a mechanistic model connecting DNA sequence organization to efficiency.

genomics↗

bioRxiv2026

Ineffectual Genomic Error Correction Under Environmental Perturbation Dynamically Regulates Mutational Supply and Robustness

Adaptive evolution depends on the supply of heritable variation, yet excessive mutation threatens viability by degrading essential molecular functions. Here, we show that this trade-off emerges naturally from the kinetic proofreading mechanism that controls replication fidelity. In our model, environmental shifts alter the optimal driving rate constant of proofreading enzymes, transiently elevating replication error rates and triggering rapid evolutionary change until a new fidelity optimum is reached. This produces alternating periods of stasis and rapid adaptation, consistent with punctuated equilibrium. We further show that coding-region length and population size jointly determine whether adaptation succeeds or mutational collapse occurs, reflecting the balance between mutation supply and error tolerance predicted by the drift-barrier principle. These results provide a molecular mechanistic basis for nearly neutral evolutionary dynamics and illustrate how genomic organization constrains long-term evolutionary resilience.

evolutionary biology↗

bioRxiv2025

Genome size and nucleotide skews as predictors of bacterial growth rate

Bacterial growth rates are constrained by genome replication, yet the role of replication kinetics in bacterial growth rates remains incompletely understood. Here, we examine if genome size, replichore organization, and nucleotide compositional asymmetry are reasonable predictors of bacterial doubling times. In free-living bacteria, both genome size and the length of the longest replichore are found to correlate positively with doubling time, pointing to an influence of replication dynamics on bacterial growth rates. Moreover, fast-growing bacteria are shown to exhibit stronger nucleotide compositional skew. Incorporating skew into the model substantially improves predictive accuracy, suggesting that compositional asymmetry in genomes may facilitate replication fork progression and thereby enhance growth rates. Based on these observations, we speculate that nucleotide skew may play a potential adaptive role in bacterial genome replication. To assess whether the observed association between genome architecture and growth rate reflects an evolutionary signature or a mechanistic link, we reconstructed ancestral states and found that the model fits ancestral traits more strongly, with predictive strength (R2) decreasing progressively along the evolutionary tree as successive speciations occur. We speculate that this association has been stronger early in bacterial evolution and became subsequently screened as organisms diversified and increased in ecological and physiological complexity.

genomics↗

bioRxiv2025

Replication rate-information storage trade-off shapes genome architecture across domains

Genome size varies widely among organisms, from compact genomes of bacteria to vast and complex genomes of eukaryotes. In this study, we theoretically identify the evolutionary pressures that may have driven this divergence in genome size. We use a parameter-free model to study genome size evolution under selection pressure to minimize replication time and maximize information storage capacity. We substantiate this choice by demonstrating a correlation between replication time and genome size using literature data. We show that bacteria tend to reduce genome size, constrained by a single replication origin, while eukaryotes expand their genomes by incorporating multiple replication origins. We propose a connection between genome size and cellular energetics, suggesting that endosymbiotic organelles, mitochondria and chloroplasts, evolutionarily regulate the number of replication origins, thereby influencing genome size in eukaryotes. We substantiate this claim by showing a correlation between organelle count and genome size data. We argue that nucleotide skews, present in nearly all genomes studied, and are used to identify replication origins, directly influence DNA unzipping kinetics, and hence its replication time. This connection enables us to derive nearly universal observations, such as Chargaffs second parity rule and replichore symmetrization, as adaptive consequences. We argue that high skews lead to faster replication, and substantiate it by showing a correlation between skews and replication speed in bacteria. The model reproduces more evolutionary genomic observations, such as a general preference for deletions over insertions across domains, and elongation and high variance of genome size under reduced selection pressure for replication rate, an integral component of C-value paradox. We highlight the possibility of regulation of the firing of latent replication origins in response to cues from the extracellular environment, leading to the regulation of cell cycle rates in multicellular eukaryotes. Significance StatementUnderstanding the forces shaping genome architecture is a long-standing challenge in evolutionary biology. Our study demonstrates that the balance between replication speed, influenced by nucleotide skews, and information storage, constrained by cellular energetics, drives the divergence in genome size between bacteria and eukaryotes. By quantifying selection pressure as the ratio of replication time to genomic information storage capacity, we show that this pressure enforces adaptive constraints, giving rise to observed features such as symmetric replichores and Chargaffs second parity rule. These insights not only help us resolve an enduring evolutionary puzzle, but also offer a unified framework linking genome organization, cellular specialization, and even potential mechanisms underlying carcinogenesis.

genomics↗

bioRxiv2025

Non-enzymatic error correction in self-replicators without extraneous energy supply

Accurate propagation of sequence information in nucleic acids is central to the evolutionary dynamics of self-replicating systems. Modern biological systems achieve high fidelity using enzymes that actively correct errors through energy-driven mechanisms. However, such complex machinery was absent under prebiotic conditions. Here, we present a theoretical model of error correction in self-replicating heteropolymers that requires neither enzymes nor an extraneous energy supply. The model relies solely on the free-energy gradient driving strand growth and requires asymmetric cooperativity -- a kinetic asymmetry known to promote unidirectional elongation. Despite its simplicity, we demonstrate that this minimal model facilitates kinetic discrimination between correct and incorrect base pair incorporations, and for specific set of parameters, reproduces the error ratio of [~] 10-4, experimentally observed in passive base selection processes. It replicates key features observed in DNA error correction, including stalling, fraying, next-nucleotide effects, and the speed-accuracy trade-off. Our results provide plausible answers for longstanding questions, such as the energy source for the enhanced base selectivity of passive DNA polymerases and the role of thermodynamics and kinetics of phosphodiester bond formation in error correction. We show that catalysis of the phosphodiester bond plays a central role in error correction, even without explicit enzymatic structural discrimination. This observation points to a plausible pathway for accurate oligomer synthesis under prebiotic conditions, driven solely by the thermodynamic gradient favoring strand elongation. More broadly, the model highlights how persistent molecular order can emerge from non-equilibrium dynamics - a central requirement for emergence of life.

biochemistry↗

bioRxiv2025

Eukaryotes evade information storage-replication rate trade-off with endosymbiont assistance leading to larger genomes

Genome length varies widely among organisms, from compact genomes of prokaryotes to vast and complex genomes of eukaryotes. In this study, we theoretically identify the evolutionary pressures that may have driven this divergence in genome length. We use a parameter-free model to study genome length evolution under selection pressure to minimize replication time and maximize information storage capacity. We show that prokaryotes tend to reduce genome length, constrained by a single replication origin, while eukaryotes expand their genomes by incorporating multiple replication origins. We propose a connection between genome length and cellular energetics, suggesting that endosymbiotic organelles, mitochondria and chloroplasts, evolutionarily regulate the number of replication origins, thereby influencing genome length in eukaryotes. We show that the above two selection pressures also lead to strict equalization of the number of purines and their corresponding base-pairing pyrimidines within a single DNA strand, known as Chagraffs second parity rule, a hitherto unexplained observation in genomes of nearly all known species. This arises from the symmetrization of replichore length, another observation that has been shown to hold across species, which our model reproduces. The model also reproduces other experimentally observed phenomena, such as a general preference for deletions over insertions, and elongation and high variance of genome lengths under reduced selection pressure for replication rate, termed the C-value paradox. We highlight the possibility of regulation of the firing of latent replication origins in response to cues from the extracellular environment leading to the regulation of cell cycle rates in multicellular eukaryotes. Significance StatementUnderstanding the forces shaping genome architecture is a long-standing challenge in evolutionary biology. Our study demonstrates that the balance between replication speed and information storage, constrained by cellular en-ergetics, drives the divergence in genome lengths between prokaryotes and eukaryotes. By quantifying selection pressure as the ratio of replication time to genomic information content, we show that this pressure enforces adaptive constraints, giving rise to features such as symmetric replichores and Chargaffs second parity rule. These insights not only help us resolve an enduring evolutionary puzzle, but also offer a unified framework linking genome organization, cellular specialization, and even potential mechanisms underlying carcinogenesis.

evolutionary biology↗

bioRxiv2019

Modeling adaptive therapy in non-muscle invasive bladder cancer

Bladder cancer is the 9th most commonly diagnosed cancer. Nearly half of patients with early stage bladder cancer treated with the immune-stimulating agent BCG have disease recurrence, while 13% progress to invasive bladder cancer. Here we explored the potential of tumor mutational heterogeneity and the role of pro- and anti-inflammatory cytokines to identify different subtypes of bladder cancer that may predict therapeutic response to BCG. Further, we used mathematical modeling of dosing strategies to infer tumor response to varying doses and time schedules f BCG administration. As a proof-of-concept, present adaptive therapy scheduling of BCG as a viable strategy to control tumor size and minimize recurrence.

cancer biology↗

bioRxiv2019

Development of a genetic toolset for the highly engineerable and metabolically versatile Acinetobacter baylyi ADP1

One primary objective of synthetic biology is to improve the sustainability of chemical manufacturing. Biological systems can utilize a variety of carbon sources, including waste streams that pose challenges to traditional chemical processing such as lignin biomass, providing opportunity for remediation and valorization of these materials. Success, however, depends on identifying microorganisms that are both metabolically versatile and engineerable. This has been a historic hindrance. Here, we leverage the facile genetics of the metabolically versatile bacterium Acinetobacter baylyi ADP1 to create easy and rapid molecular cloning workflows, a promoter library, ribosomal binding site (RBS) variants, and an unprecedented number of bacterial chromosomal protein expression sites and variants. Moreover, we demonstrate the utility of these tools by examining ADP1s catabolic repression regulation, creating a strain with improved potential for lignin bioprocessing. Taken together, this work establishes ADP1 as an ideal host for a variety of sustainability and synthetic biology applications.

synthetic biology↗

bioRxiv2018

Multimodal interferometric imaging of nanoscale structure and macromolecular motion uncovers UV induced cellular paroxysm

We present a multimodal label-free interferometric imaging platform for measuring intracellular nanoscale structure and macromolecular dynamics in living cells with a sensitivity to macromolecules as small as 20nm and millisecond temporal resolution. We validate this system by pairing experimental measurements of nanosphere phantoms with a novel interferometric theory. Applying this system in vitro, we explore changes in higher-order chromatin structure and dynamics that occur due to cellular fixation, stem cell differentiation, and ultraviolet (UV) light irradiation. Finally, we discover a new phenomenon, cellular paroxysm, a near-instantaneous, synchronous burst of motion that occurs early in the process of UV induced cell death. Given this platforms ability to obtain nanoscale sensitive, millisecond resolved information within live cells without concerns of photobleaching, it has the potential to answer a broad range of critical biological questions about macromolecular behavior in live cells, particularly about the relationship between cellular structure and function.

biophysics↗

bioRxiv2018

Preservation of cellular nano-architecture by the process of chemical fixation for nanopathology

Transformation in chromatin organization is one of the most universal markers of carcinogenesis. Microscale chromatin alterations have been a staple of histopathological diagnosis of neoplasia, and nanoscale alterations have emerged as a promising marker for cancer prognostication and the detection of predysplastic changes. While numerous methods have been developed to detect these alterations, most methods for sample preparation remain largely validated via conventional microscopy, and have not been examined with nanoscale sensitive imaging techniques. For these nanoscale sensitive techniques to become standard of care screening tools, new histological protocols must be developed that preserve nanoscale information. Partial Wave Spectroscopic (PWS) microscopy has recently emerged as a novel imaging technique sensitive to length scales ranging between 20 and 200 nanometers. As a label-free, high-throughput, and non-invasive imaging technique, PWS microscopy offers many advantages for risk stratification of early cancer, and is an ideal tool to quantify structural information during sample preparation. Therefore, in this work we applied PWS microscopy to systematically evaluate the effects of cytological preparation on the nanoscales changes of chromatin using two cell line models: Hela cells differentially treated with daunorubicin and TP53 differentially mutated ovarian carcinoma cells. Notably, we show that existing cytological preparation can be modified in order to maintain clinically relevant nanoscopic differences, paving the way for the emerging field of nanopathology.

biophysics↗