Interplay between gene expression and gene architecture as a consequence of gene and genome duplications: evidence from metabolic genes of Arabidopsis thaliana.

Gene and genome duplications have been widespread during the evolution of flowering plant which resulted in the increment of biological complexity as well as creation of plasticity of a genome helping the species to adapt to changing environments. Duplicated genes with higher evolutionary rates can act as a mechanism of generating novel functions in secondary metabolism. In this study, we explored duplication as a potential factor governing the expression heterogeneity and gene architecture of Primary Metabolic Genes (PMGs) and Secondary Metabolic Genes (SMGs) of Arabidopsis thaliana. It is remarkable that different types of duplication processes controlled gene expression and tissue specificity differently in PMGs and SMGs. A complex relationship exists between gene architecture and expression patterns of primary and secondary metabolic genes. Our study reflects, expression heterogeneity and gene structure variation of primary and secondary metabolism in Arabidopsis thaliana are partly results of duplication events of different origins. Our study suggests that duplication has differential effect on PMGs and SMGs regarding expression pattern by controlling gene structure, epigenetic modifications, multifunctionality and subcellular compartmentalization. This study provides an insight into the evolution of metabolism in plants in the light of gene and genome scale duplication. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-022-01188-2.

Chaperone client proteins evolve slower than non-client proteins.

Chaperones are important molecular machinery that assists proteins to attain their native three-dimensional structure crucial for function. Earlier studies using experimental evolution showed that chaperones impose a relaxation of sequence constraints on their "client" proteins, which may lead to the fixation of slightly deleterious mutations on the latter. However, we hypothesized that such a phenomenon might be harmful to the organism in a natural physiological condition. In this study, we investigated the evolutionary rates of chaperone client and non-client proteins in five model organisms from both prokaryotic and eukaryotic lineages. Our study reveals a slower evolutionary rate of chaperone client proteins in all five organisms. Additionally, the slower folding rate and lower aggregation propensity of chaperone client proteins reveal that the chaperone may play an essential role in rescuing the slightly disadvantageous effects due to random mutations and subsequent protein misfolding. However, the fixation of such mutations is less likely to be selected in the natural population.

Exploring the major cross-talking edges of competitive endogenous RNA networks in human Chronic and Acute Myeloid Leukemia.

BACKGROUND: Human Chronic and Acute Myeloid Leukemia are myeloproliferative disorders in myeloid lineage of blood cells characterized by accumulation of aberrant white blood cells. In cancer, the anomalous transcriptome includes deregulated expression of non-coding RNAs in conjunction with protein-coding mRNAs in human genome. The coding or non-coding RNA transcripts harboring miRNA-binding sites can converse with and regulate each other by explicitly contending for a limited pool of shared miRNAs and act as competitive endogenous RNAs (ceRNAs). An unifying hypothesis attributing 'modulation of expression of transcripts' in this fashion had been defined as 'competitive endogenous RNA hypothesis'. Network built with ceRNAs evidently offers a platform to elucidate complex regulatory interactions at post-transcriptional level in human cancers. METHODS: Contemplating cancers of human myeloid lineage we constructed ceRNA networks for CML and AML coding and non-coding repertoire utilizing patient sample data. Through functional enrichment analysis we selected the significant functional modules for transcripts being differentially expressed in Blastic phases of each cancer types with respect to Normal. After retrieving free energy of binding and duplex formation of shared miRNAs on ceRNAs, we performed statistical averaging of energy values over the ensemble of populations considering cellular system as in canonical (Iso-thermal) situation. RESULTS AND CONCLUSIONS: We aimed to shed light on 'Sibling Rivalry' in ceRNA partners from the perspective of statistical thermodynamics, identified major cross-talking tracks and ceRNAs influencing transcripts concerned in myeloid cancer systems. GENERAL SIGNIFICANCE: Insights into ceRNA-regulation will shed light on progression and prognosis of human Chronic and Acute Myeloid Leukemia.

Evolutionary rate heterogeneity between multi- and single-interface hubs across human housekeeping and tissue-specific protein interaction network: Insights from proteins' and its partners' properties.

Integrating gene expression into protein-protein interaction network (PPIN) leads to the construction of tissue-specific (TS) and housekeeping (HK) sub-networks, with distinctive TS- and HK-hubs. All such hub proteins are divided into multi-interface (MI) hubs and single-interface (SI) hubs, where MI hubs evolve slower than SI hubs. Here we explored the evolutionary rate difference between MI and SI proteins within TS- and HK-PPIN and observed that this difference is present only in TS, but not in HK-class. Next, we explored whether proteins' own properties or its partners' properties are more influential in such evolutionary discrepancy. Statistical analyses revealed that this evolutionary rate correlates negatively with protein's own properties like expression level, miRNA count, conformational diversity and functional properties and with its partners' properties like protein disorder and tissue expression similarity. Moreover, partial correlation and regression analysis revealed that both proteins' and its partners' properties have independent effects on protein evolutionary rate.

The role of introns in the conservation of the metabolic genes of Arabidopsis thaliana.

In Arabidopsis thaliana, primary metabolic genes (PMGs) are more evolutionarily conserved and intron-rich than secondary metabolic genes. We observed that PMGs are more primitive and pan-taxonomically persistent as compared to secondary (SMGs) and non-metabolic genes (NMGs). This difference in primitiveness and persistence is primarily correlated with intron number and is independent of gene expression level. We propose a twofold explanation behind higher intron enrichment in PMGs. Firstly, introns might increase protein versatility amongst PMGs through alternative splicing, providing selective advantage of PMGs and making them more persistent across diverse plant taxa. Also, multifunctional PMGs may acquire functional domains by increasing the intronic burden. Additionally, single nucleotide polymorphisms (SNPs) accumulate at a higher rate in introns as compared to exons. Moreover, a strong negative correlation between cumulative exonic SNPs density and intron number indicates that introns may protect the exonic regions against the deleterious effect of these mutations, making them more conserved.

Systematic Analyses and Prediction of Human Drug Side Effect Associated Proteins from the Perspective of Protein Evolution.

Identification of various factors involved in adverse drug reactions in target proteins to develop therapeutic drugs with minimal/no side effect is very important. In this context, we have performed a comparative evolutionary rate analyses between the genes exhibiting drug side-effect(s) (SET) and genes showing no side effect (NSET) with an aim to increase the prediction accuracy of SET/NSET proteins using evolutionary rate determinants. We found that SET proteins are more conserved than the NSET proteins. The rates of evolution between SET and NSET protein primarily depend upon their noncomplex (protein complex association number = 0) forming nature, phylogenetic age, multifunctionality, membrane localization, and transmembrane helix content irrespective of their essentiality, total druggability (total number of drugs/target), m-RNA expression level, and tissue expression breadth. We also introduced two novel terms-killer druggability (number of drugs with killing side effect(s)/target), essential druggability (number of drugs targeting essential proteins/target) to explain the evolutionary rate variation between SET and NSET proteins. Interestingly, we noticed that SET proteins are younger than NSET proteins and multifunctional younger SET proteins are candidates of acquiring killing side effects. We provide evidence that higher killer druggability, multifunctionality, and transmembrane helices support the conservation of SET proteins over NSET proteins in spite of their recent origin. By employing all these entities, our Support Vector Machine model predicts human SET/NSET proteins to a high degree of accuracy (∼86%).

Codon degeneracy and amino acid abundance influence the measures of codon usage bias: improved Nc (N̂c ) and ENCprime (N̂'c ) measures.

Effective number of codons (N^c) and its variant N^'c (effective number of codons prime) are the two widely used methods for measuring unequal usage of synonymous codons in coding sequences, known as the codon usage bias (CUB). The mathematical formula used in calculating N^c and N^'c values is giving inappropriate measures of CUB in case of low abundance of amino acids. In addition, the magnitude of error also varies according to codon degeneracy. In this study, a modified formula for N^c and N^'c has been developed to measure the CUB more accurately. Online implementations of the modified formula are available in the web portal at http://agnigarh.tezu.ernet.in/~ssankar/cub.php.

Deciphering the cross-talking of human competitive endogenous RNAs in K562 chronic myelogenous leukemia cell line.

Chronic myelogenous leukemia (CML) is a myeloproliferative disorder characterized by increased proliferation or abnormal accumulation of granulocytic cell line without the depletion of their capacity to differentiate. A reciprocal chromosomal translocation proceeding to the 'Philadelphia chromosome', involving the ABL proto-oncogene and BCR gene residing on Chromosome 9 and 22 respectively, is observed to be attributed to CML pathogenesis. Recent studies have been unraveling the crucial role of genomic 'dark matter' or the non-coding repertoire in cancer initiation and progression. The intricate cross-talk between competitive endogenous RNAs (ceRNAs) provides a scaffold to systematically functionalize the miRNA response element harboring non-coding RNAs and incorporate them with the protein-coding RNA dimension in complex ceRNA networks. This network of coding and non-coding transcriptome linked by shared miRNAs evidently offers a platform to elucidate the complex regulatory interactions at the post-transcriptional level in human cancers. In this context, analyzing CML, from the perspective of the ceRNA hypothesis, surely craves intensive attention and a comprehensive discussion. Here, we performed RNA-seq data analysis to retrieve Lymphoblastoid and CML coding as well as non-coding repertoire and constructed a ceRNA network for the CML cell line, considering the non-cancer lymphoblastoid cell line as the control. We investigated if any alteration exists in the ceRNA landscape of the transcripts which are exhibiting differential expression across the two cell lines and observed that the major ceRNA regulators vary in cancer network when compared with the Lymphoblastoid network. The top ranked significant functional modules in the ceRNA network display cancer associated attributes and reveal putative regulators in CML pathogenesis.

Insights into the dN/dS ratio heterogeneity between brain specific genes and widely expressed genes in species of different complexity.

In mammals, it has long been suggested that brain-specific genes (BSGs) and widely expressed genes (WEGs) have seemingly lower dN/dS ratio than any other gene sets. However, to what extent these genes differ in their dN/dS ratio has still remained controversial. Here, we have revealed lower dN/dS ratio of BSGs than WEGs in human-mouse, human-orangutan, human-chimpanzee and mouse-rat orthologous pair. The significance level of dN/dS ratio difference indicates a trend of decreasing difference as complexity of compared pairs increases. Further studies with the human-mouse pair revealed that, removal of the duplicated genes from both the dataset has nullified this difference which dictates a vital role of duplicated genes in governing the selection pressure. Conclusively, higher paralog number, expression level, and longer regulatory region length of BSGs allow fewer nucleotide substitutions within them. Our results show for the first time to our knowledge lower dN/dS ratio of BSGs than WEGs.

Overlapping genes: A significant genomic correlate of prokaryotic growth rates.

Elucidating the genomic features influencing prokaryotic growth rates has always been a study of interest. Previously, it was observed that overlapping genes (OGs) play a crucial role in the prokaryotic genome size reduction. This study is focused to explore whether OGs act as a potential correlate of prokaryotic growth rates. For this purpose, we compiled a dataset of 25 archaeal and 117 eubacterial genomes and analyzed the inter-correlation between the proportion of overlapping regions in these genomes with their growth rates. Here, we observed that the proportion of overlapping region holds a significant negative correlation with generation time in archaeal domain, whereas no correlation was observed in the eubacterial domain. However, after masking the effect of tRNA, rRNA multiplicity and environmental diversity, OGs show an independent effect over growth rates in the eubacterial domain as well as in the archaeal domain. Moreover, the influence of OGs on prokaryotic growth rates provides different delineations in archaeal and eubacterial domains. In archaea, both long overlap frequency (LOF) and short overlap frequency (SOF) influence the growth rates by increasing the degree of operonization. On the contrary, in the case of bacteria, neither SOF nor LOF plays any significant role in achieving faster growth rates.

Exploring the evolutionary rate differences between human disease and non-disease genes.

Comparisons of evolutionary features between human disease and non-disease genes have a wide implication to understand the genetic basis of human disease genes. However, it has not yet been resolved whether disease genes evolve at slower or faster rate than the non-disease genes. To resolve this controversy, here we integrated human disease genes from several databases and compared their protein evolutionary rates with non-disease genes in both housekeeping and tissue-specific group. We noticed that in tissue specific group, disease genes evolve significantly at a slower rate than non-disease genes. However, we found no significant difference in evolutionary rates between disease and non-disease genes in housekeeping group. Tissue specific disease genes have a higher protein complex number, elevated gene expression level and are also associated with conserve biological processes. Finally, our regression analysis suggested that protein complex number followed by protein multifunctionality independently modulates the evolutionary rate of human disease genes.

Evolutionary Rate Heterogeneity of Primary and Secondary Metabolic Pathway Genes in Arabidopsis thaliana.

Primary metabolism is essential to plants for growth and development, and secondary metabolism helps plants to interact with the environment. Many plant metabolites are industrially important. These metabolites are produced by plants through complex metabolic pathways. Lack of knowledge about these pathways is hindering the successful breeding practices for these metabolites. For a better knowledge of the metabolism in plants as a whole, evolutionary rate variation of primary and secondary metabolic pathway genes is a prerequisite. In this study, evolutionary rate variation of primary and secondary metabolic pathway genes has been analyzed in the model plant Arabidopsis thaliana. Primary metabolic pathway genes were found to be more conserved than secondary metabolic pathway genes. Several factors such as gene structure, expression level, tissue specificity, multifunctionality, and domain number are the key factors behind this evolutionary rate variation. This study will help to better understand the evolutionary dynamics of plant metabolism.

Elucidating evolutionary features and functional implications of orphan genes in Leishmania major.

Orphan genes are protein coding genes that lack recognizable homologs in other organisms. These genes were reported to comprise a considerable fraction of coding regions in all sequenced genomes and thought to be allied with organism's lineage-specific traits. However, their evolutionary persistence and functional significance still remain elusive. Due to lack of homologs with the host genome and for their probable lineage-specific functional roles, orphan gene product of pathogenic protozoan might be considered as the possible therapeutic targets. Leishmania major is an important parasitic protozoan of the genus Leishmania that is associated with the disease cutaneous leishmaniasis. Therefore, evolutionary and functional characterization of orphan genes in this organism may help in understanding the factors prevailing pathogen evolution and parasitic adaptation. In this study, we systematically identified orphan genes of L. major and employed several in silico analyses for understanding their evolutionary and functional attributes. To trace the signatures of molecular evolution, we compared their evolutionary rate with non-orphan genes. In agreement with prior observations, here we noticed that orphan genes evolve at a higher rate as compared to non-orphan genes. Lower sequence conservation of orphan genes was previously attributed solely due to their younger gene age. However, here we observed that together with gene age, a number of genomic (like expression level, GC content, variation in codon usage) and proteomic factors (like protein length, intrinsic disorder content, hydropathicity) could independently modulate their evolutionary rate. We considered the interplay of all these factors and analyzed their relative contribution on protein evolutionary rate by regression analysis. On the functional level, we observed that orphan genes are associated with regulatory, growth factor and transport related processes. Moreover, these genes were found to be enriched with various types of interaction and trafficking motifs, implying their possible involvement in host-parasite interactions. Thus, our comprehensive analysis of L. major orphan genes provided evidence for their extensive roles in host-pathogen interactions and virulence.

Overlapping genes: a new strategy of thermophilic stress tolerance in prokaryotes.

Overlapping genes (OGs) draw the focus of recent day's research. However, the significance of OGs in prokaryotic genomes remained unexplored. As an adaptation to high temperature, thermophiles were shown to eliminate their intergenic regions. Therefore, it could be possible that prokaryotes would increase their OG content to adapt to high temperature. To test this hypothesis, we carried out a comparative study on OG frequency of 256 prokaryotic genomes comprising both thermophiles and non-thermophiles. It was found that thermophiles exhibit higher frequency of overlapping genes than non-thermophiles. Moreover, overlap frequency was found to correlate with optimal growth temperature (OGT) in prokaryotes. Long overlap frequency was found to hold a positive correlation with OGT resulting in an abundance of long overlaps in thermophiles compared to non-thermophiles. On the other hand, short overlap (1-4 nucleotides) frequency (SOF) did not yield any direct correlation with OGT. However, the correlation of SOF with CAIavg (extent of variation of codon usage bias measured as the mean of codon adaptation index of all genes in a given genome) and IG% (proportion of intergenic regions) indicate that they might upregulate the aforementioned factors (CAIavg and IG%) which are already known to be vital forces for thermophilic adaptation. From these evidences, we propose that the OG content bears a strong link to thermophily. Long overlaps are important for their genome compaction and short overlaps are important to uphold high CAIavg. Our findings will surely help in better understanding of the significance of overlapping gene content in prokaryotic genomes.

Insights into the miRNA regulations in human disease genes.

BACKGROUND: MicroRNAs are a class of short non-coding RNAs derived from either cellular or viral transcripts that act post-transcriptionally to regulate mRNA stability and translation. In recent days, increasing numbers of miRNAs have been shown to be involved in the development and progression of a variety of diseases. We, therefore, intend to enumerate miRNA targets in several known disease classes to explore the degree of miRNA regulations on them which is unexplored till date. RESULTS: Here, we noticed that miRNA hits in cancer genes are remarkably higher than other diseases in human. Our observation suggests that UTRs and the transcript length of cancer related genes have a significant contribution in higher susceptibility to miRNA regulation. Moreover, gene duplication, mRNA stability, AREScores and evolutionary rate were likely to have implications for more miRNA targeting on cancer genes. Consequently, the regression analysis have confirmed that the AREScores plays most important role in detecting miRNA targets on disease genes. Interestingly, we observed that epigenetic modifications like CpG methylation and histone modification are less effective than miRNA regulations in controlling the gene expression of cancer genes. CONCLUSIONS: The intrinsic properties of cancer genes studied here, for higher miRNA targeting will enhance the knowledge on cancer gene regulation.

Elucidating the genotype-phenotype relationships and network perturbations of human shared and specific disease genes from an evolutionary perspective.

To date, numerous studies have been attempted to determine the extent of variation in evolutionary rates between human disease and nondisease (ND) genes. In our present study, we have considered human autosomal monogenic (Mendelian) disease genes, which were classified into two groups according to the number of phenotypic defects, that is, specific disease (SPD) gene (one gene: one defect) and shared disease (SHD) gene (one gene: multiple defects). Here, we have compared the evolutionary rates of these two groups of genes, that is, SPD genes and SHD genes with respect to ND genes. We observed that the average evolutionary rates are slow in SHD group, intermediate in SPD group, and fast in ND group. Group-to-group evolutionary rate differences remain statistically significant regardless of their gene expression levels and number of defects. We demonstrated that disease genes are under strong selective constraint if they emerge through edgetic perturbation or drug-induced perturbation of the interactome network, show tissue-restricted expression, and are involved in transmembrane transport. Among all the factors, our regression analyses interestingly suggest the independent effects of 1) drug-induced perturbation and 2) the interaction term of expression breadth and transmembrane transport on protein evolutionary rates. We reasoned that the drug-induced network disruption is a combination of several edgetic perturbations and, thus, has more severe effect on gene phenotypes.

GC-made protein disorder sheds new light on vertebrate evolution.

At the emergence of endothermic vertebrates, GC rich regions of the ectothermic ancestral genomes underwent a significant GC increase. Such an increase was previously postulated to increase thermodynamic and structural stability of proteins through selective increase of protein hydrophobicity. Here, we found that, increase in GC content promotes a higher content of disorder promoting amino acid in endothermic vertebrates proteins and that the increase in hydrophobicity is mainly due to a higher content of the small disorder promoting amino acid alanine. In endothermic vertebrates, prevalence of disordered residues was found to promote functional diversity of proteins encoded by GC rich genes. Higher fraction of disordered residues in this group of proteins was also found to minimize their aggregation tendency. Thus, we propose that the GC transition has favored disordered residues to promote functional diversity in GC rich genes, and to protect them against functional loss by protein misfolding.

Prevalent structural disorder carries signature of prokaryotic adaptation to oxic atmosphere.

Microbes have adopted efficient mechanisms to contend with environmental changes. The emergence of oxygen was a major event that led to an abrupt change in Earth's atmosphere. To adjust with this shift in environmental condition ancient microbes must have undergone several modifications. Although some proteomic and genomic attributes were proposed to facilitate survival of microorganisms in the presence of oxygen, the process of adaptation still remains elusive. Recent studies have focused that intrinsically disordered proteins play crucial roles in adaptation to a wide range of ecological conditions. Therefore, it is likely that disordered proteins could also play indispensable roles in microbial adaptation to the aerobic environment. To test this hypothesis we measured the disorder content of 679 prokaryotes from four oxygen requirement groups. Our result revealed that aerobic proteomes are endowed with the highest protein disorder followed by facultative microbes. Minimal disorder was observed in anaerobic and microaerophilic microbes with no significant difference in their disorder content. Considering all the potential confounding factors that can modulate protein disorder, here we established that the high protein disorder in aerobic microbe is not a by-product of adaptation to any other selective pressure. On the functional level, we found that the high disorder in aerobic proteomes has been utilized for processes that are important for their aerobic lifestyle. Moreover, aerobic proteomes were found to be enriched with disordered binding sites and to contain transcription factors with high disorder propensity. Based on our results, here we proposed that the high protein disorder is an adaptive opportunity for aerobic microbes to fit with the genomic and functional complexities of the aerobic lifestyle.

Pseudogenes and their composers: delving in the 'debris' of human genome.

Pseudogenes, the nonfunctional homologs of functional genes and thus exemplified as 'genomic fossils' provide intriguing snapshots of the evolutionary history of human genome. These defunct copies generally arise by retrotransposition or duplication followed by various genetic disablements. In this study, focusing on human pseudogenes and their functional homologues we describe their characteristic features and relevance to protein sequence evolution. We recapitulate that pseudogenes harbor disease-causing degenerative sequence variations in conjunction with the immense disease gene association of their progenitors. Furthermore, we also discuss the issue of functional resurrection and the potentiality observed in some pseudogenes to regulate their functional counterparts.

Evolutionary rate heterogeneity of core and attachment proteins in yeast protein complexes.

In general, proteins do not work alone; they form macromolecular complexes to play fundamental roles in diverse cellular functions. On the basis of their iterative clustering procedure and frequency of occurrence in the macromolecular complexes, the protein subunits have been categorized as core and attachment. Core protein subunits are the main functional elements, whereas attachment proteins act as modifiers or activators in protein complexes. In this article, using the current data set of yeast protein complexes, we found that core proteins are evolving at a faster rate than attachment proteins in spite of their functional importance. Interestingly, our investigation revealed that attachment proteins are present in a higher number of macromolecular complexes than core proteins. We also observed that the protein complex number (defined as the number of protein complexes in which a protein subunit belongs) has a stronger influence on gene/protein essentiality than multifunctionality. Finally, our results suggest that the observed differences in the rates of protein evolution between core and attachment proteins are due to differences in protein complex number and expression level. Moreover, we conclude that proteins which are present in higher numbers of macromolecular complexes enhance their overall expression level by increasing their transcription rate as well as translation rate, and thus the protein complex number imposes a strong selection pressure on the evolution of yeast proteome.