Biology subjects

Govindaraju, A.

Publications and source records attributed to Govindaraju, A..

4 recordsLinked to original sources

bioRxiv2026

Local GPCR density tips the balance of μ-opioid receptor trafficking

The extent to which local GPCR surface density governs engagement of downstream signaling and trafficking pathways remains unclear. Using single-particle tracking of the -opioid receptor (MOR), we show that receptor density differentially regulates G protein signaling and GRK2/3-{beta}-arrestin-dependent receptor trafficking. At low surface density, MORs activate G proteins but fail to enter clathrin-coated structures despite the presence of endogenous GRK2/3 and {beta}-arrestin. Increasing MOR density, co-expressing other class A GPCRs, or elevating GRK2 or {beta}-arrestin abundance rescues agonist-induced MOR trafficking. In contrast, the class B GPCR V2R blocks MOR trafficking at both low and high MOR densities. These results support a model in which increasing class A GPCR density, despite worsening effector-to-receptor stoichiometry, promotes trafficking by forming an affinity matrix that enables reversible GRK2/3 and {beta}-arrestin interactions to be productively used by neighboring receptors in a density-dependent manner, whereas class B GPCRs sequester {beta}-arrestin and block trafficking.

biophysics↗

bioRxiv2023

Characterization of a phyllosphere methylotroph community isolated with lanthanides

Lanthanides have emerged as important metal cofactors for biological processes. Lanthanide-associated metabolisms are well-studied in leaf symbiont methylotrophic bacteria, which utilize reduced one-carbon compounds such as methanol for growth. Yet, the importance of lanthanides in plant-microbe interactions and on microbial physiology and colonization in plants remains poorly understood. To investigate this, 344 pink-pigmented facultative methylotrophs were isolated from soybean leaves by selecting for bacteria capable of methanol oxidation with lanthanide cofactors, but none were obligately lanthanide-dependent. Phylogenetic analyses revealed that all strains were nearly identical to each other and are part of the extorquens clade of Methylobacterium, despite variability in genome and plasmid sizes. Strain-specific identification was enabled by the higher resolution provided with rpoB compared to 16S rRNA as marker genes. Despite the low strain-level diversity, the metabolic capabilities of the collection diverged greatly. Strains encoding identical lanthanide-dependent alcohol dehydrogenases displayed significantly different growth rates and/or final ODs from each other on alcohols in the presence and absence of lanthanides. Several strains also lacked well-characterized lanthanide-associated genes thought to be important for phyllosphere colonization. Additionally, 3% of our isolates were capable of growth on sugars and 23% were capable of growth on aromatic acids, substantially expanding the range of substrates utilized by Methylobacterium extorquens in the phyllosphere. Our findings suggest that the expansion of metabolic capabilities, as well as differential usage of lanthanides and their influence on metabolism, among closely related strains point to evolution of niche partitioning strategies to promote colonization of the phyllosphere. ImportanceLanthanide metals have long been appreciated for their role in technology, but the recent identification of lanthanides as cofactors in methylotrophic metabolism has expanded the role of lanthanides into biology. In the phyllosphere, methylotrophs are some of the most abundant bacteria found on leaf surfaces, where lanthanide concentrations are sufficiently high to support their growth. Yet, the extent to which lanthanides influence methylotrophic metabolism in the phyllosphere remains unknown. Here, we characterize a methylotrophic enrichment isolated from the phyllosphere in a lanthanide-dependent manner. We have identified (1) closely related strains with identical lanthanide-dependent enzymes that exhibit different growth on alcohols in the presence of lanthanides, (2) resilient strains lacking lanthanide-associated genes thought to be important for phyllosphere colonization, and (3) many strains capable of metabolisms that were thought to be rare within this clade. Overall, our isolates serve as a model community to interrogate how lanthanides differentially influence methylotrophic physiology.

microbiology↗

bioRxiv2021

Methylothon: a versatile course-based high school research experience in microbiology and bioinformatics-- with pink bacteria

Methylothon is an inquiry-based high school learning module in microbial ecology, molecular biology, and bioinformatics that centers around pink-pigmented plant-associated methylotrophic bacteria. Here we present an overview of the modules learning goals, describe course resources (available for public use on http://methylothon.com), and relate lessons learned from adapting Methylothon for remote learning during the pandemic in spring of 2021. The original in-person version of the module allows students to isolate their own strains of methylotrophic bacteria from plants they sample from the environment, to identify these using PCR, sequencing, and phylogenetic analysis, and to contribute their strains to original research in a university lab. The adapted version strengthens the focus on bioinformatics and increases its flexibility and accessibility by making the lab portion optional and adopting free web-based tools. Student feedback and graded assignments from Spring 2021 revealed that the lesson was especially effective at introducing the concepts of BLAST and phylogenetic trees, and that students valued and felt inspired by the opportunity to conduct hands-on work and to participate in community science.

scientific communication and education↗

bioRxiv2018

Phototrophic lactate utilization by Rhodopseudomonas palustris is stimulated by co-utilization with additional substrates

The phototrophic purple nonsulfur bacterium Rhodopseudomonas palustris is known for its metabolic versatility and is of interest for various industrial and environmental applications. Despite decades of research on R. palustris growth under diverse conditions, patterns of R. palustris growth and carbon utilization with mixtures of carbon substrates remain largely unknown. R. palustris readily utilizes most short chain organic acids but cannot readily use lactate as a sole carbon source. Here we investigated the influence of mixed-substrate utilization on phototrophic lactate consumption by R. palustris. We found that lactate was simultaneously utilized with a variety of other organic acids and glycerol in time frames that were insufficient for R. palustris growth on lactate alone. Thus, lactate utilization by R. palustris was expedited by its co-utilization with additional substrates. Separately, experiments using carbon pairs that did not contain lactate revealed acetate-mediated inhibition of glycerol utilization in R. palustris. This inhibition was specific to the acetate-glycerol pair, as R. palustris simultaneously utilized acetate or glycerol when either was paired with succinate or lactate. Overall, our results demonstrate that (i) R. palustris commonly employs simultaneous mixed-substrate utilization, (ii) mixed-substrate utilization expands the spectrum of readily utilized organic acids in this species, and (iii) R. palustris has the capacity to exert carbon catabolite control in a substrate-specific manner.\n\nIMPORTANCEBacterial carbon source utilization is frequently assessed using cultures provided single carbon sources. However, the utilization of carbon mixtures by bacteria (i.e., mixed-substrate utilization) is of both fundamental and practical importance; it is central to bacterial physiology and ecology, and it influences the utility of bacteria as biotechnology. Here we investigated mixed-substrate utilization by the model organism Rhodopseudomonas palustris. Using mixtures of organic acids and glycerol, we show that R. palustris exhibits an expanded range of usable carbon substrates when provided in mixtures. Specifically, co-utilization enabled the prompt consumption of lactate, a substrate that is otherwise not readily used by R. palustris. Additionally, we found that R. palustris utilizes acetate and glycerol sequentially, revealing that this species has the capacity to use some substrates in a preferential order. These results provide insights into R. palustris physiology that will aid the use of R. palustris for industrial and commercial applications.

microbiology↗