Anusha Gopalakrishnan

• Doctor of Philosophy
• Research Associate at Duke University Medical Center

Pathogens have been a strong driving force for natural selection. Therefore, understanding how human genetic differences impact infection-related cellular traits can mechanistically link genetic variation to disease susceptibility. Here we report the Hi-HOST Phenome Project (H2P2): a catalog of cellular genome-wide association studies (GWAS) compri...

Genome-wide association studies (GWAS) have identified thousands of genetic variants associated with disease. To facilitate moving from associations to disease mechanisms, we leveraged the role of pathogens in shaping human evolution with the Hi-HOST Phenome Project (H2P2): a catalog of cellular GWAS comprised of 79 phenotypes in response to 8 path...

In sexually reproducing organisms, meiosis is an essential step responsible for generation of haploid gametes from diploid somatic cells. The quest for understanding regulatory mechanisms of meiotic recombination in Plasmodium led to identification of a gene encoding a protein that contains 11 copies of C2H2 zinc fingers (ZnF). Reverse genetic appr...

Chlamydia trachomatis is a leading cause of genital and ocular infections for which no vaccine exists. Upon entry into host cells, C. trachomatis resides within a membrane-bound compartment-the inclusion-and secretes inclusion membrane proteins (Incs) that are thought to modulate the host-bacterium interface. To expand our understanding of Inc func...

Artemisinin-based combination therapy (ACT) is the recommended first-line treatment for Plasmodium falciparum malaria. It has been suggested that the cytotoxic effect of artemisinin is mediated by free radicals followed by the alkylation of P. falciparum proteins. The endoperoxide bridge, the active moiety of artemisinin derivatives, is cleaved in...

We report the clinical and analytical performance of an isothermal thermophilic helicase-dependent amplification assay for blood Plasmodium parasite detection and species-level identification. The assay amplifies the 18S rRNA gene fragment of all Plasmodium species and uses a species-specific probe and a pan-malarial probe to definitively identify...

Unlabelled: The bacterial RecA protein and its eukaryotic homologue Rad51 play a central role in the homologous DNA strand exchange reaction during recombination and DNA repair. Previously, our lab has shown that PfRad51, the Plasmodium falciparum homologue of Rad51, exhibited ATPase activity and promoted DNA strand exchange in vitro. In this stud...

(A) Role of CaCl2 (0.5 mM) in PfRad51-mediated SSE. (B) SSE showing various intermediate products when the gel was run for a shorter time period. Aliquots were collected at time points (min) indicated above each lane and quenched with stop solution, and products were revealed on 1% TAE agarose gel, followed by EtBr staining. Lds, linear double-stra...

Reverse transcriptase-PCR of pfgrp. Parasite cultures used for MMS induction studies for real-time PCR (results in Fig. 5) were independently verified for the absence of any trace of contaminating gDNA. Cultures were synchronized by sorbitol treatment, and cDNA was synthesized using total RNA purified from rings (lanes 1 to 3), trophozoites (lanes...

Alignment of various domains (A, B, and C, represented in Fig. 1A) of PfRad54 with ScRad54 using BLAST analysis showing percent identity, positives, and E values. Download

Percent homology of the amino acid sequences of Plasmodium recombination molecules with those of yeast, human, Leishmania, Trypanosome, and Cryptosporidium. Percent homology of PfRPA1S is also shown with Cryptosporidium and Toxoplasma, the only other two apicomplexans that contain the short form of RPA1. PfRad51, a key molecule in homologous recomb...

Reverse transcriptase-PCR analysis in the presence and absence of RT for pfrad51. Parasite cultures were synchronized by sorbitol treatment, and cDNA was synthesized using RNA from MMS-induced (0.05%) rings, trophozoites, and schizonts (lanes 1). In parallel, samples from 0.05% MMS-treated samples were processed in the absence of reverse transcript...

PfRPA1S negatively impacts the function of PfRPA1L. (A) In the absence of PfRPA1S, PfRPA1L serves the function of SSB and PfRad51-mediated SSE is catalyzed. (B) Addition of PfRPA1S reduces PfRad51-mediated SSE. (C) Excess of PfRPA1S negatively impacts the function of PfRPA1L, and no SSE occurs. Download

Nucleotide sequences of primers used for gene-specific amplification for expression. Engineered restriction enzyme sites (underlined) were added to each primer to facilitate directional cloning into the pRsetA vector.

Nucleotide sequences of primers used for real-time PCR analysis.

Malaria threatens millions of people annually and is a burden to human health and economic development. Unfortunately in terms of disease control, no effective vaccines are available and the efficacy of treatment is limited by drug resistance. Genetic manipulation in Plasmodium falciparum is hampered due to the absence of robust methods for genetic...

SynonymsComet assays; Damage and repair; Double-strand breaks; Drug resistance; Methyl methanesulfonate ; Mismatch repair; MutS and MutL; Rad51Definition/SummaryDNA damage arises from exposure of DNA to damaging agents. Damage response pathway is activated upon chromosomal perturbations. DNA repair processes correct DNA damage and restore genomic i...

Apicomplexans comprise some of the most life threatening parasites infecting human and livestock and includes Plasmodium and Toxoplasma, the causative agents of malaria and toxoplasmosis respectively, in humans as well as Neospora caninum (abortion in livestock, neosporosis in dogs), Cryptosporidium (Diarrheal cryptosporidiosis and opportunistic in...

Identification of promoter elements responsible for regulation of gene expression has been hampered by the AT richness of P. falciparum intergenic regions. Nested deletions of histidine-rich protein 3 (hrp3) promoter suggested the presence of a multipartite ring-specific element. Linker scanning (LS) of this ring-specific promoter showed that the a...

Over 80% of Plasmodium falciparum genes are developmentally regulated during the parasite’s life cycle with most genes expressed in a “just in time” fashion. However, the molecular mechanisms of gene regulation are still poorly understood. Analysis of P. falciparum genome shows that the parasite appears to encode relatively few transcription factor...

The asexual blood stage of Plasmodium falciparum in the human host is comprised of morphologically distinct ring, trophozoite and schizont stages, each of which possesses a distinct pattern of gene expression. Episomal promoter recombination has been recently reported in malaria parasites. We aim to investigate the nature of this process, and its r...

The asexual blood stage of Plasmodium falciparum is comprised of morphologically distinct ring, trophozoite and schizont stages. Each of these developmental stages possesses a distinct pattern of gene expression. Regulation of P. falciparum gene expression is thought to occur, at least in part, at the promoter level. Previously, we have found that...