Biology subjects

Ramkumar, S.

Publications and source records attributed to Ramkumar, S..

5 recordsLinked to original sources

bioRxiv2026

pH-dependent anti-TfR1 NANOBODY(R) molecules deliver efficacious oligonucleotide payloads to muscle and CNS tissues

The blood-brain barrier (BBB) is a highly selective, semi-permeable border of endothelial cells that prevents solutes and therapeutic agents in systemic circulation from passively crossing into the central nervous system (CNS) parenchyma. The Transferrin receptor 1 (TfR1) endocytosis pathway for iron homeostasis is one of the most well-characterized strategies for therapeutic delivery across the BBB. The work presented here showcases the discovery of novel anti-TfR1 NANOBODY(R) shuttles. The identified anti-TfR1 NANOBODY(R) molecules display cross-reactivity and pH-dependent binding to human, cynomolgus (cyno), and mouse TfR1. Structural data further explain and support the underlying mechanism of this pH-dependent binding. These anti-TfR1 NANOBODY(R) molecules were successfully conjugated to both short-interfering RNA (siRNA) and antisense oligonucleotide (ASO) tool payloads. anti-TfR1 NANOBODY(R)-siRNA conjugates can induce up to 60% knockdown of the target mRNA transcript in skeletal muscle up to two weeks post a single IV dose in mice and up to 35-40% at four weeks post dose. Furthermore, extending the half-life of the anti-TfR1 NANOBODY(R)-ASO shuttles enhances heart, sciatic nerve, and brain exposure and enables up to 30-60% target knockdown in different CNS cell types. Altogether, these results highlight important features for the development of anti-TfR1 shuttles for the purpose of downregulating target mRNA transcripts in muscle and CNS for a variety of neurologic and neuromuscular indications.

neuroscience↗

bioRxiv2023

Functional characterization of five triterpene synthases through De-novo assembly and transcriptome analysis of Euphorbia grantii and Euphorbia tirucalli

Euphorbia grantii and Euphorbia tirucalli known to synthesize diverse triterpenes including euphol and tirucallol. These two triterpenes known to possess potent anti-cancer, anti-bacterial, and anti-fungal properties along with various other biological activities. In this study, De-novo assembly and comparative transcriptome analysis of leaf and stem tissues of E. tirucalli and E. grantii were carried out to identify thirteen triterpene synthases from 1,40,227 in correlation with the metabolic profiling. Comparative transcriptome analysis indicated that EutTTS4 and EutTTS5 genes which encodes for euphol/tirucallol and tirucallol synthase were highly expressed in leaf and stem tissue. The genes which encodes -amyrin synthase (EutTTS1) and lupeol synthase (EutTTS2) were characterized by overexpressing them in YPH499 yeast strain. We have developed using hem1 and erg7 knock yeast strain of lanosterol deficient yeast (TMBL17) and used for over expression of friedelin synthase (EutTTS3), and two novel triterpenes synthases such as euphol/tirucallol synthase (EutTTS4) and tirucallol synthase (EutTTS5). These results are very useful in large scale production of triterpenes by genomic integration of respective triterpene synthases in TMBL yeast strain developed in this study. Significance StatementWe have functionally characterized triterpene synthases from E. tirucalli and E. grantii and developed a hem1 and erg7 knock out of lanosterol deficient yeast (TMBL17) for the large-scale production of triterpene and triterpene related products.

molecular biology↗

bioRxiv2021

Disturbed retinoid metabolism upon loss of rlbp1a impairs cone function and leads to subretinal lipid deposits and photoreceptor degeneration in the zebrafish retina

The RLBP1 gene encodes the 36 kDa cellular retinaldehyde binding protein, CRALBP, a soluble retinoid carrier, in the visual cycle of the eyes. Mutations in RLBP1 are associated with recessively inherited clinical phenotypes, including Bothnia dystrophy, retinitis pigmentosa, retinitis punctata albescens, fundus albipunctatus, and Newfoundland rod-cone dystrophy. However, the etiology of these retinal disorders is not well understood. Here, we generated homologous zebrafish models to bridge this knowledge gap. Duplication of the rlbp1 gene in zebrafish and cell-specific expression of the paralogs rlbp1a in the retinal pigment epithelium and rlbp1b in Muller glial cells allowed us to create intrinsically cell type-specific knockout fish lines. Using rlbp1a and rlbp1b single and double mutants, we investigated the pathological effects on visual function. Our analyses revealed that rlbp1a was essential for cone photoreceptor function and chromophore metabolism in the fish eyes. rlbp1a mutant fish displayed reduced chromophore levels and attenuated cone photoreceptor responses to light stimuli. They accumulated 11-cis and all-trans-retinyl esters which displayed as enlarged lipid droplets in the RPE reminiscent of the subretinal yellow-white lesions in patients with RLBP1 mutations. During aging, these fish developed retinal thinning and cone and rod photoreceptor dystrophy. In contrast, rlbp1b mutants did not display impaired vision. The double mutant essentially replicated the phenotype of the rlbp1a single mutant. Together, our study showed that the rlbp1a zebrafish mutant recapitulated many features of human blinding diseases caused by RLBP1 mutations and provided novel insights into the pathways for chromophore regeneration of cone photoreceptors. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC="FIGDIR/small/448986v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@bd3de5org.highwire.dtl.DTLVardef@2a9420org.highwire.dtl.DTLVardef@162e7f0org.highwire.dtl.DTLVardef@697ee6_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2019

Cardiac valve regeneration in adult zebrafish: importance of TGF-β signaling in new tissue formation

Cardiac valve disease can lead to severe cardiac dysfunction and is thus a frequent cause of morbidity and mortality. Its main treatment is valve replacement, which is currently greatly limited by the poor recellularization and tissue formation potential of the implanted valves. As we still lack suitable animal models to identify modulators of these processes, here we used the adult zebrafish and found that, upon valve decellularization, they initiate a striking regenerative program that leads to the formation of new functional valves. After injury, endothelial and kidney marrow-derived cells undergo cell cycle re-entry and differentiate into new extracellular matrix-secreting valve cells. The Transforming Growth Factor beta (TGF{beta}) signaling pathway promotes this process by enhancing progenitor cell proliferation as well as valve cell differentiation. These findings reveal a key role for TGF{beta} signaling in valve regeneration and also establish the zebrafish as a model to identify and test factors promoting valve recellularization and growth.

developmental biology↗

bioRxiv2018

Role of biofilm-specific antibiotic resistance genes PA0756-0757, PA5033 and PA2070 in Pseudomonas aeruginosa

To evaluate the presence of biofilm-specific antibiotic-resistant genes, PA0756-0757, PA5033 and PA2070 in Pseudomonas aeruginosa isolated from clinical samples in Tamil Nadu. For this cross-sectional study, 24 clinical isolates (included pus, urine, wound, and blood) were collected from two diagnostic centers in Chennai from May 2015 to February 2016. Biofilm formation was assessed using microtiter dish biofilm formation assay and minimal inhibitory concentration (MIC) and minimal bactericidal concentrations (MBC) were determined for planktonic and biofilm cells (MBC assay). Further, PCR amplification of biofilm-specific antibiotic resistance genes PA0756-0757, PA5033 and PA2070 were performed. Biofilm formation was found to be moderate/strong in 16 strains. MBC for planktonic cells showed that 4, 7, 10 and 14 strains were susceptible to gentamicin, ciprofloxacin, meropenem and colistin respectively. In MBC assay for biofilm cells (MBC-B), all the 16 biofilm producing strains were resistant to ciprofloxacin and gentamicin whereas nine and four were resistant to meropenem, and colistin respectively. The biofilm-specific antibiotic-resistant genes PA0756-0757 was found in 10 strains, 6 strains with PA5033 and 9 strains with PA2070 that were found to be resistant phenotypically. This study highlighted the importance of biofilm-specific antibiotic resistance genes PA0756-0757, PA5033, and PA2070 in biofilm-forming P. aeruginosa.

microbiology↗