Dynamics of DNA Methylation in Recent Human and Great Ape Evolution

DNA methylation is an epigenetic modification involved in regulatory processes such as cell differentiation during development, X-chromosome inactivation, genomic imprinting and susceptibility to complex disease. However, the dynamics of DNA methylation changes between humans and their closest relatives are still poorly understood. We performed a comparative analysis of CpG methylation patterns between 9 humans and 23 primate samples including all species of great apes (chimpanzee, bonobo, gorilla and orangutan) using Illumina Methylation450 bead arrays. Our analysis identified ∼800 genes with significantly altered methylation patterns among the great apes, including ∼170 genes with a methylation pattern unique to human. Some of these are known to be involved in developmental and neurological features, suggesting that epigenetic changes have been frequent during recent human and primate evolution. We identified a significant positive relationship between the rate of coding variation and alterations of methylation at the promoter level, indicative of co-occurrence between evolution of protein sequence and gene regulation. In contrast, and supporting the idea that many phenotypic differences between humans and great apes are not due to amino acid differences, our analysis also identified 184 genes that are perfectly conserved at protein level between human and chimpanzee, yet show significant epigenetic differences between these two species. We conclude that epigenetic alterations are an important force during primate evolution and have been under-explored in evolutionary comparative genomics.     

Genetic Diversity Among Cold-Tolerant Fluorescent Pseudomonas Isolates from Indian Himalayas and Their Characterization for Biocontrol and Plant Growth-Promoting Activities

In Uttarakhand, the Organic State of India, where soils in most farming situations are deficient in nutrients and loss of crops due to soil- and seed-borne pathogens is rampant, use of native plant growth-promoting rhizobacteria (PGPRs) possessing biocontrol (BC) activities holds promise. In view of this, 600 native cold-tolerant rhizospheric bacterial isolates were collected from Uttarakhand Himalayas, of which 336 were confirmed as fluorescent Pseudomonas spp. On the basis of specific biochemical tests, these were characterized into three major groups: P. fluorescens (308 isolates), P. aeruginosa (20 isolates), and P. putida (8 isolates). Most of the isolates could grow at 8°C after 12 h of incubation, confirming their cold tolerance. In vitro biocontrol assays revealed that of 336 isolates, 74 were antagonistic to Rhizoctonia solani and 91 to Fusarium solani, the two major pathogens associated with root-rot complex in vegetables widespread in the region. Simultaneously, good HCN producers (33 isolates), siderophore producers (80 isolates), and P solubilizers (49 isolates) were also identified, which could increase the biocontrol and plant growth-promoting efficacies of the putative PGPRs. Among the different species and biovars, P. fluorescens biovar-I had the maximum number of potential isolates with BC and plant growth-promoting (PGP) activities. In French bean, under polyhouse and field conditions, five isolates (Pf-173, Pf-193, Pf-547, Pf-551, and Pf-572) showed good BC and PGP activities as up to 93% reduction in root rot was achieved. A combination of all five isolates was found to be best with respect to BC and PGP activities. In a set of 59 fluorescent Pseudomonas isolates, RAPD-PCR analysis, using three random oligodecamer primers, revealed high diversity and formed ten distinct clusters, corresponding to the host of origin (annual or perennial) or habitat (farming situations) of the isolates. The amount of diversity revealed in the set of fluorescent Pseudomonas isolates could represent enormous diversity that exists in the wild that could be exploited for improved BC and PGP activities of the PGPRs. For the first time, this study led to a large-scale characterization and repositioning of fluorescent pseudomonads from the Indian Himalayas.     

The Effects of Salinity on Nitrogen Fixation and Trehalose Metabolism in Mycorrhizal Cajanus cajan (L.) Millsp. Plants

Arbuscular mycorrhizal (AM) fungi exist widely in natural ecosystems as well as in salt-affected soils and are considered suitable candidates for bio-amelioration of saline soils. Plants respond to salinity by accumulating sugars and other low-molecular-weight compatible solutes. One such compound is trehalose, which has been found to play an important role as a stress protectant. The aim of the present investigation was to study interactions between an AM fungus and salinity stress on growth, nitrogen fixation, and trehalose metabolism in Cajanus cajan (L.) Millsp. (pigeonpea). Two genotypes [Sel 85N (salt-tolerant) and ICP 13997 (salt-sensitive)] were subjected to saline treatments with and without mycorrhizal inoculations. Salinity reduced plant biomass (shoot and root) in both genotypes and resulted in a decline in shoot-to-root ratio (SRR); however, a smaller decline was observed in Sel 85N than in ICP 13997. AM colonization was reduced with increasing salinity levels but mycorrhizal responsiveness (MR) increased. Genotypic variability in nitrogen fixation and trehalose metabolism in response to salinity and mycorrhization was observed. An increment in nodule number was accompanied by a reduction in dry mass. Subsequently, nodular activity (leghemoglobin, acetylene-reduction activity [ARA], nitrogen content) was reduced under soil salinity, which was more profound in ICP 13997 than in Sel 85N. The symbiotic association with Glomus mosseae led to significant improvement in plant dry mass and nitrogen-fixing potential of nodules under salt stress. Salinity led to an increase in trehalose-6-P synthetase (TPS) and trehalose-6-P phosphatase (TPP) activities resulting in increased trehalose content in nodules, which was accompanied by inhibition of trehalose catabolism (trehalase activity). AM plants had lower trehalase activity under saline and nonsaline conditions. Thus, a symbiotic relationship between plant roots and G. mosseae might have resulted in salinity tolerance in a genotype-dependent manner.     

The Rv1651c-encoded PE-PGRS30 protein expressed in Mycobacterium smegmatis exhibits polar localization and modulates its growth profile

Sequencing analysis of the complete genome of Mycobacterium tuberculosis (Mtb) H37Rv resulted in the identification of a novel multigene, the PE family of genes. The genes of the largest PE_PGRS subfamily of the PE family are mainly restricted to pathogenic mycobacteria, and their exact role in the biology of Mtb is not clearly understood. Based on their sequence homology, PE_PGRS proteins were initially thought to serve common functions. However, studies on individual proteins reveal that the individual proteins of this subfamily could be performing several unrelated tasks. In the present study, we investigated the function of PE_PGRS30 by expressing it in Mycobacterium smegmatis. PE_PGRS30 expression in M. smegmatis resulted in phenotypic changes with altered colony morphology and growth profile. The recombinant PE_PGRS30 showed polar localization and was found to be associated with the cell wall of M. smegmatis. Thus, the present study suggests that the prolonged lag phase of growth caused by the PE_PGRS30 may, in part, contribute to the latency of Mtb.     

Application of a novel bacterial consortium for mineralization of sulphonated aromatic amines

A novel bacterial consortium (TJ-2) for mineralization of aromatic amines resulting from decolorization of azo dyes was developed. Three bacterial strains were identified as Pseudomonas pseudoalcaligenes (TJ-21,EU072476), Pseudomonas citronellolis (TJ-22,EU072477) and Pseudomonas testosterone (TJ-23,EU072477) by 16S rRNA gene sequence analysis. Aromatic amine mineralization under aerobic conditions was observed to be significantly higher with the consortium as compared to pure strains indicating complementary interactions among these strains. It was observed that more than 90% mineralization of aromatic amines was achieved within 18 h for different initial aromatic amines concentrations. It was also observed that aromatic amine mineralization depends upon the structure of aromatic amine. Para- and meta-hydroxy substituted aromatic amine were easily mineralized as compared to ortho-substituted which undergoes autoxidation when exposed to oxygen. The consortium was capable of mineralizing other aromatic amines, thus, conferring the possibility of application of TJ-2 for the treatment of industrial wastewaters containing aromatic amines.     

Identification and Functional Analysis of the Primary Pantothenate Transporter,PfPAT, of the Human Malaria Parasite Plasmodium falciparum

The human malaria parasite Plasmodium falciparum is absolutely dependent on the acquisition of host pantothenate for its development within human erythrocytes. Although the biochemical properties of this transport have been characterized, the molecular identity of the parasite-encoded pantothenate transporter remains unknown. Here we report the identification and functional characterization of the first protozoan pantothenate transporter, PfPAT, from P. falciparum. We show using cell biological, biochemical, and genetic analyses that this transporter is localized to the parasite plasma membrane and plays an essential role in parasite intraerythrocytic development. We have targeted PfPAT to the yeast plasma membrane and showed that the transporter complements the growth defect of the yeast fen2Δ pantothenate transporter-deficient mutant and mediates the entry of the fungicide drug, fenpropimorph. Our studies in P. falciparum revealed that fenpropimorph inhibits the intraerythrocytic development of both chloroquine- and pyrimethamine-resistant P. falciparum strains with potency equal or better than that of currently available pantothenate analogs. The essential function of PfPAT and its ability to deliver both pantothenate and fenpropimorph makes it an attractive target for the development and delivery of new classes of antimalarial drugs.     

Cell‐specific effects on surface α7 nicotinic receptor expression revealed by over‐expression and knockdown of rat RIC3 protein

We tested whether surface α7 nicotinic acetylcholine receptor expression is dependent on an endogenous chaperone named Resistance to Inhibitors of Cholinesterase 3 (RIC3) by comparing RIC3 protein in rat GH4C1 and human SH‐EP1 cells, which express strikingly different surface receptor levels following α7 transfection. Cloned rat RIC3 exists in at least two isoforms because of an ambiguous splice site between exons 4 and 5. Both rat isoforms permit surface α7 expression in SH‐EP1 and human embryonic kidney (HEK) cells measured by α‐bungarotoxin binding. Contrary to expectations, endogenous RIC3 protein expression determined by immunoblots did not differ between untransfected GH4C1 or SH‐EP1 cells. siRNA against rat RIC3 exon 4 and shRNA against exons 2, 5 and 6 knocked down transfected rat RIC3 expression in SH‐EP1 cells and simultaneously blocked toxin binding. However, no RNAi construct blocked binding when co‐transfected with α7 into GH4C1 cells. shRNA against rat exons 2 and 5 knocked down rat RIC3 protein transfected into GH4C1 cells with a time course suggesting a protein half‐life of a few days. These results suggest GH4C1 cells may possess unknown chaperone(s) allowing high surface α7 expression in the absence of known RIC3 splice variants.     

A draft genome sequence of the pulse crop chickpea (Cicer arietinum L.)

Cicer arietinum L. (chickpea) is the third most important food legume crop. We have generated the draft sequence of a desi‐type chickpea genome using next‐generation sequencing platforms, bacterial artificial chromosome end sequences and a genetic map. The 520‐Mb assembly covers 70% of the predicted 740‐Mb genome length, and more than 80% of the gene space. Genome analysis predicts the presence of 27 571 genes and 210 Mb as repeat elements. The gene expression analysis performed using 274 million RNA‐Seq reads identified several tissue‐specific and stress‐responsive genes. Although segmental duplicated blocks are observed, the chickpea genome does not exhibit any indication of recent whole‐genome duplication. Nucleotide diversity analysis provides an assessment of a narrow genetic base within the chickpea cultivars. We have developed a resource for genetic markers by comparing the genome sequences of one wild and three cultivated chickpea genotypes. The draft genome sequence is expected to facilitate genetic enhancement and breeding to develop improved chickpea varieties.