The Ancient Phosphatidylinositol 3-Kinase Signaling System Is a Master Regulator of Energy and Carbon Metabolism in Algae

Ramanan, Rishiram,Tran, Quynh-Giao,Cho, Dae-Hyun,Jung, Jae-Eun,Kim, Byung-Hyuk,Shin, Sang-Yoon,Choi, Sae-Hae,Liu, Kwang-Hyeon,Kim, Dae-Soo,Lee, Seon-Jin,Crespo, José,L.,Lee, Hee-Gu,Oh, Hee-Mock American Society of Plant Biologists 2018 PLANT PHYSIOLOGY - Vol.177 No.3

<P>Phosphatidylinositol 3-kinase signaling influences biofuel yields in algae by regulating membrane lipid hydrolysis, lipogenesis, tricarboxylic acid cycle, and mitochondrial ATP synthesis.</P><P>Algae undergo a complete metabolic transformation under stress by arresting cell growth, inducing autophagy and hyper-accumulating biofuel precursors such as triacylglycerols and starch. However, the regulatory mechanisms behind this stress-induced transformation are still unclear. Here, we use biochemical, mutational, and “omics” approaches to demonstrate that PI3K signaling mediates the homeostasis of energy molecules and influences carbon metabolism in algae. In <I>Chlamydomonas reinhardtii</I>, the inhibition and knockdown (KD) of algal class III PI3K led to significantly decreased cell growth, altered cell morphology, and higher lipid and starch contents. Lipid profiling of wild-type and PI3K KD lines showed significantly reduced membrane lipid breakdown under nitrogen starvation (−N) in the KD. RNA-seq and network analyses showed that under −N conditions, the KD line carried out lipogenesis rather than lipid hydrolysis by initiating de novo fatty acid biosynthesis, which was supported by tricarboxylic acid cycle down-regulation and via acetyl-CoA synthesis from glycolysis. Remarkably, autophagic responses did not have primacy over inositide signaling in algae, unlike in mammals and vascular plants. The mutant displayed a fundamental shift in intracellular energy flux, analogous to that in tumor cells. The high free fatty acid levels and reduced mitochondrial ATP generation led to decreased cell viability. These results indicate that the PI3K signal transduction pathway is the metabolic gatekeeper restraining biofuel yields, thus maintaining fitness and viability under stress in algae. This study demonstrates the existence of homeostasis between starch and lipid synthesis controlled by lipid signaling in algae and expands our understanding of such processes, with biotechnological and evolutionary implications.</P>

Influence of CO₂ concentration on carbon concentrating mechanisms in cyanobacteria and green algae: a proteomic approach

Ramanan, Rishiram,Vinayagamoorthy, Nadimuthu,Sivanesan, Saravana Devi,Kannan, Krishnamurthi,Chakrabarti, Tapan The Korean Society of Phycology 2012 ALGAE Vol.27 No.4

Carbon concentrating mechanisms play a vital role in photosynthesis in microalgae and cyanobacteria especially in the proper functioning of Rubisco and assimilation of carbon via the Calvin cycle. This study evaluates the role of carbon dioxide on carbon concentrating mechanism (CCM) in a cynaobacteria, Spirulina platensis and a microalga, Chlorella sp. 786. The study organisms were grown in both atmospheric (control sample, 0.035%) and high (exposed sample, 10%) $CO_2$ concentrations. Second dimension (2D) electrophoresis revealed a huge difference in the protein profiles of both organisms suggesting the induction of CCM related proteins in the sample maintained at atmospheric $CO_2$ concentration and the repression of CCM related proteins in the sample maintained at 10% $CO_2$. Liquid chromatography-mass spectroscopy analysis revealed the presence of two important $C_i$ transporter proteins in the control sample of S. platensis, namely ferredoxin-$NADP^+$ reductase and ATP binding cassette (ABC) transport system protein. These proteins were only expressed in the control sample and were downregulated or not expressed at all in the exposed sample. Consequently, this study conclusively proves that CCMs are only inducted at low $CO_2$ concentrations and are not functional at high $CO_2$ concentration.

Algae-bacteria interactions: Evolution, ecology and emerging applications

Ramanan, R.,Kim, B.H.,Cho, D.H.,Oh, H.M.,Kim, H.S. Pergamon Press ; Elsevier Science Ltd 2016 BIOTECHNOLOGY ADVANCES Vol.34 No.1

<P>Algae and bacteria have coexisted ever since the early stages of evolution. This coevolution has revolutionized life on earth in many aspects. Algae and bacteria together influence ecosystems as varied as deep seas to lichens and represent all conceivable modes of interactions from mutualism to parasitism. Several studies have shown that algae and bacteria synergistically affect each other's physiology and metabolism, a classic case being algae-roseobacter interaction. These interactions are ubiquitous and define the primary productivity in most ecosystems. In recent years, algae have received much attention for industrial exploitation but their interaction with bacteria is often considered a contamination during commercialization. A few recent studies have shown that bacteria not only enhance algal growth but also help in flocculation, both essential processes in algal biotechnology. Hence, there is a need to understand these interactions from an evolutionary and ecological standpoint, and integrate this understanding for industrial use. Here we reflect on the diversity of such relationships and their associated mechanisms, as well as the habitats that they mutually influence. This review also outlines the role of these interactions in key evolutionary events such as endosymbiosis, besides their ecological role in biogeochemical cycles. Finally, we focus on extending such studies on algal-bacterial interactions to various environmental and bio-technological applications. (C) 2015 The Authors. Published by Elsevier Inc This is an open access article under the CC BY-NC-ND license.</P>

Phycosphere bacterial diversity in green algae reveals an apparent similarity across habitats

Ramanan, Rishiram,Kang, Zion,Kim, Byung-Hyuk,Cho, Dae-Hyun,Jin, Long,Oh, Hee-Mock,Kim, Hee-Sik Elsevier 2015 Algal research Vol.8 No.-

<P><B>Abstract</B></P> <P>Phytoplankton and bacteria play the foremost role in primary production and often act in unison in biogeochemical cycling. Studies conducted so far are inconclusive on species specificity of phycosphere bacteria as the overarching function of specific clades of algae-associated bacteria, for instance <I>Roseobacter</I> in sulfur cycling, is widely held. In this study, we attempt to demonstrate the diversity of phycosphere bacteria in phylogenetically divergent unialgal green algae from vastly different environmental samples like soil, freshwater, marine, and wastewater with diatom and cyanobacteria as an outgroup. Diversity analyses using Differential Gel Gradient Electrophoresis (DGGE) revealed the predominant presence of bacteria belonging to Bacteroidetes phylum (46% of all strains). 454 pyrosequencing of selected strains from different habitats not only confirmed the presence of Bacteroidetes (33.1% of total reads) but also revealed the presence of bacteria belonging to α-Proteobacteria (52.6%), all in close association with their host. Majority of those symbiotic bacteria have been classified as Plant Growth Promoting Bacteria (PGPB) including prominent Sphingomonads and Rhizobacter. Results suggest that although host algae might encourage species specific interactions, specific functional traits are prerequisite for proximal adhesion in nutrient-rich phycosphere. While Bacteroidetes is known to have significant role in nutrient cycling through degradation of plant and algal macromolecules and for its attached growth, PGPB have proven symbiosis with plants and the overwhelming presence of these bacteria in green algae points to possible co-evolution.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Phycosphere bacterial diversity of 11 algal strains from different habitats by DGGE. </LI> <LI> DGGE & pyrosequencing revealed the dominance of α-Proteobacteria and Bacteroidetes. </LI> <LI> Algal-bacterial interactions might not be species specific. </LI> <LI> Algae might prefer certain bacterial clades with specific functional traits. </LI> <LI> Results of this study has ecological and biotechnological implications. </LI> </UL> </P>

Response of flame thickness and propagation speed under intense turbulence in spatially developing lean premixed methane–air jet flames

Sankaran, Ramanan,Hawkes, Evatt R.,Yoo, Chun Sang,Chen, Jacqueline H. Elsevier 2015 Combustion and Flame Vol.162 No.9

<P><B>Abstract</B></P> <P>Direct numerical simulations of three-dimensional spatially-developing turbulent Bunsen flames were performed at three different turbulence intensities. The simulations were performed using a reduced methane–air chemical mechanism which was specifically tailored for the lean premixed conditions simulated here. A planar-jet turbulent Bunsen flame configuration was used in which turbulent preheated methane–air mixture at 0.7 equivalence ratio issued through a central jet and was surrounded by a hot laminar coflow of burned products. The turbulence characteristics at the jet inflow were selected such that combustion occured in the thin reaction zones (TRZ) regime. At the lowest turbulence intensity, the conditions fall on the boundary between the TRZ regime and the corrugated flamelet regime, and progressively moved further into the TRZ regime by increasing the turbulent intensity. The data from the three simulations was analyzed to understand the effect of turbulent stirring on the flame structure and thickness. Statistical analysis of the data showed that the thermal preheat layer of the flame was thickened due to the action of turbulence, but the reaction zone was not significantly affected. A global and local analysis of the burning velocity of the flame was performed to compare the different flames. Detailed statistical averages of the flame speed were also obtained to study the spatial dependence of displacement speed and its correlation to strain rate and curvature.</P>

Cytomegalovirus Infections in Solid Organ Transplantation: A Review

Poornima Ramanan,Raymund R Razonable 대한감염학회 2013 Infection and Chemotherapy Vol.45 No.3

Cytomegalovirus (CMV) continues to have a tremendous impact in solid organ transplantation despite remarkable advances in its diagnosis, prevention and treatment. It can affect allograft function and increase patient morbidity and mortality through a number of direct and indirect effects. Patients may develop asymptomatic viremia, CMV syndrome or tissue-invasive disease. Late-onset CMV disease continues to be a major problem in high-risk patients after completion of antiviral prophylaxis. Emerging data suggests that immunologic monitoring may be useful in predicting the risk of late onset CMV disease. There is now increasing interest in the development of an effective vaccine for prevention. Novel antiviral drugs with unique mechanisms of action and lesser toxicity are being developed. Viral load quantification is now undergoing standardization, and this will permit the generation of clinically relevant viral thresholds for the management of patients. This article provides a brief overview of the contemporary epidemiology, clinical presentation, diagnosis, prevention and treatment of CMV infection in solid organ transplant recipients.

Purification and Characterization of a Novel Plant-type Carbonic Anhydrase from Bacillus subtilis

Rishiram Ramanan,Krishnamurthi Kannan,Nadimuthu Vinayagamoorthy,Saravana Devi Sivanesan,Kunga Mohan Ramkumar,Tapan Chakrabarti 한국생물공학회 2009 Biotechnology and Bioprocess Engineering Vol.14 No.1

Carbonic anhydrase enzyme, one of the fastest known enzymes, remains largely unexplored in prokaryotes when compared to its mammalian counterparts despite its ubiquity. In this study, the enzyme has been purified from Bacillus subtilis SA3 using sequential Sephadex G-75 chromatography, DEAE cellulose chromatography, and sepharose-4B-L-tyrosine-sulphanilamide affinity chromatography and characterized to provide additional insights into its properties. The apparent molecular mass of carbonic anhydrase obtained by SDS-PAGE was found to be approximately 37 kDa. Isoelectric focusing of the purified enzyme revealed an isoelectric point (pI) of around 6.1 when compared with marker. The presence of metal ions such as Zn2+, Co2+, Cu2+, Fe3+, Mg2+, and anion SO4- increased enzyme activity while strong inhibition was observed in the presence of Hg2+, Cl-, HCO3-, and metal chelator EDTA. The optimum pH and temperature for the enzyme were found to be 8.3 and 37℃, respectively. Enzyme kinetics with p-nitrophenyl acetate as substrate at pH 8.3 and 37℃ determined the Vmax and Km values of the enzyme to be 714.28 μmol/mg protein/min and 9.09 mM, respectively. The Ki value for acetazolamide was 0.22 mM, compared to 0.099 mM for sulphanilamide. The results from N-terminal amino acid sequencing imply the purified protein is a putative beta-carbonic anhydrase with close similarities to CAs from plants, microorganisms.

Influence of CO2 concentration on carbon concentrating mechanisms in cyanobacteria and green algae: a proteomic approach

Rishiram Ramanan,Nadimuthu Vinayagamoorthy,Saravana Devi Sivanesan,Krishnamurthi Kannan,Tapan Chakrabarti 한국조류학회I 2012 ALGAE Vol.27 No.4

Carbon concentrating mechanisms play a vital role in photosynthesis in microalgae and cyanobacteria especially in the proper functioning of Rubisco and assimilation of carbon via the Calvin cycle. This study evaluates the role of carbon dioxide on carbon concentrating mechanism (CCM) in a cynaobacteria, Spirulina platensis and a microalga, Chlorella sp. 786. The study organisms were grown in both atmospheric (control sample, 0.035%) and high (exposed sample, 10%)CO2 concentrations. Second dimension (2D) electrophoresis revealed a huge difference in the protein profiles of both organisms suggesting the induction of CCM related proteins in the sample maintained at atmospheric CO2 concentration and the repression of CCM related proteins in the sample maintained at 10% CO2. Liquid chromatography-mass spectroscopy analysis revealed the presence of two important Ci transporter proteins in the control sample of S. platensis,namely ferredoxin-NADP+ reductase and ATP binding cassette (ABC) transport system protein. These proteins were only expressed in the control sample and were downregulated or not expressed at all in the exposed sample. Consequently,this study conclusively proves that CCMs are only inducted at low CO2 concentrations and are not functional at high CO2concentration.

The Performance of a Glass Bead Shaking Technique for the Disruption of Escherichia coli Cells

Ramakrishnan Nagasundara Ramanan,Tau Chuan Ling,Arbakariya B. Ariff 한국생물공학회 2008 Biotechnology and Bioprocess Engineering Vol.13 No.5

The efficacy of a simple laboratory method for cell disruption based on the shaking of glass beads on a rotary shaker was assessed in this study, via measurements of the release of total protein and interferon-2b from E. coli. The op-timum conditions for cell disruption were detected after 30 min of shaking in Tris-HCl buffer (pH 8) at 300 rpm with 1.5 g of glass beads (diameter: 0.5 mm) per mL of cell suspension volume. Three test runs were conducted under the above conditions and the maximum average protein release values were determined as 3.048, 3.564, and 3.015 mg/mL, respectively. The amount of protein release was comparable to the amount of protein release in ultrasonica-tion and glass bead vortexing procedures. The amount of interferon-2b release in the ultrasonication, glass bead vortexing, and glass bead shaking trials were 240, 172, and 201 ng/mL, respectively. This method was shown to process between 1 and 10 mL of sample volume in a 50 mL Falcon tube without a great deal of deviation, and was able to handle in excess of 60 samples simultaneously.

Intra-operative fractures in primary total knee arthroplasty - a systematic review

Purudappa Prabhudev Prasad,Ramanan Sruthi Priyavadhana,Tripathy Sujit Kumar,Varatharaj Sushrruti,Mounasamy Varatharaj,Sambandam Senthil Nathan 대한슬관절학회 2020 대한슬관절학회지 Vol.32 No.-

Background: One of the rare complications of primary total knee arthroplasty is intra-operative fracture. Intraoperative fracture during revision knee arthroplasty has been well-documented but there is limited literature on fractures occurring during primary knee arthroplasty. We conducted a systematic review of the literature to compare and contrast the various studies to clearly define the predisposing factors, incidence, and characteristics of the fracture itself and to arrive at a consensus on the management and prevention of intra-operative fractures during primary knee arthroplasty. Methods: The PubMed/Medline, Cochrane, Scopus and Embase databases were searched using keywords “intraoperative fracture”, “distal femoral fracture”, “tibial fracture”, “patella fracture” and “primary total knee arthroplasty”. A total of 158 articles were retrieved and after further filtration and exclusion processing, 10 articles that evaluated intra-operative fractures in primary total knee arthroplasty were included for the review. Results: The reported incidence of intra-operative fractures varied from 0.2% to 4.4%. A higher incidence in female patients with a male to female ratio of 0.4 was reported. Posterior stabilized (PS) total knee arthroplasty was associated with higher risk of intra-operative femoral fractures by many authors in this review. Timing of occurrence and location of the intra-operative fractures can vary widely, with femoral fractures occurring more commonly during bone preparation, trialing and impaction of the final implant and tibial fractures occurring during preparation for the tibial keel and impaction of the tibial component. Conclusions: Intra-operative fractures during primary total knee arthroplasty are rare with higher risk associated with osteoporosis, rheumatoid arthritis, advanced age, female gender, chronic steroid use, metabolic bone disorders, PS type of femoral implant and difficult surgical exposure of the knee joint due to severe deformities. A plethora of management options have been utilized according to surgeon preference. Standard principles of fracture fixation and arthroplasty principles should be followed to achieve stable internal fixation and any unstable fracture site should be bypassed with the utilization of stemmed components. Satisfactory radiographic and functional outcome can be expected with appropriate treatment.