Biology Commons^™

Genetic And Physical Interactions Between The Organellar Mechanosensitive Ion Channel Homologs Msl1, Msl2, And Msl3 Reveal A Role For Inter-Organellar Communication In Plant Development, Josephine S. Lee, Margaret E. Wilson, Ryan A. Richardson, Elizabeth S. Haswell

Biology Faculty Research

Plant development requires communication on many levels, including between cells and between organelles within a cell. For example, mitochondria and plastids have been proposed to be sensors of environmental stress and to coordinate their re- sponses. Here we present evidence for communication between mitochondria and chloroplasts during leaf and root development, based on genetic and physical inter- actions between three Mechanosensitive channel of Small conductance-Like (MSL) proteins from Arabidopsis thaliana. MSL proteins are Arabidopsis homologs of the bac- terial Mechanosensitive channel of Small conductance (MscS), which relieves cellular osmotic pressure to protect against lysis during hypoosmotic shock. MSL1 localizes to …

The Yeast Mitochondrial Proteins Rcf1 And Rcf2 Support The Enzymology Of The Cytochrome C Oxidase Complex And Generation Of The Proton Motive Force, Vera Strogolova, Ngoc H. Hoang, Jonathan Hosler, Rosemary A. Stuart

Biological Sciences Faculty Research and Publications

The yeast mitochondrial proteins Rcf1 and Rcf2 are associated with a subpopulation of the cytochrome bc₁–cytochrome c oxidase supercomplex and have been proposed to play a role in the assembly and/or modulation of the activity of the cytochrome c oxidase (complex IV, CIV). Yeast mutants deficient in either Rcf1 or Rcf2 proteins can use aerobic respiration–based metabolism for growth, but the absence of both proteins results in a strong growth defect. In this study, using assorted biochemical and biophysical analyses of Rcf1/Rcf2 single and double null-mutant yeast cells and mitochondria, we further explored how Rcf1 and Rcf2 support …

Congruent Phylogenetic Relationships Of Melaphidina Aphids (Aphididae: Eriosomatinae: Fordini) According To Nuclear And Mitochondrial Dna Data With Taxonomic Implications On Generic Limits, Zhumei Ren, Carol D. Von Dohlen, A. J. Harris, Rebecca B. Dikow, Xu Su, Jun Wen

Biology Faculty Publications

Melaphidina aphids (Rhus-gall aphids; Eriosomatinae: Fordini) comprise five genera from eastern Asia and one monotypic genus from eastern North America. Melaphidina are unique in feeding on plant species of Rhus subgenus Rhus (Anacardiaceae), on which they form galls during the summer. The phylogenetic relationships among some species of Melaphidina aphids remain controversial. In this study, we sought to resolve the backbone phylogeny of Melaphidina aphids by sampling 15 accessions representing all six genera, all species, and all subspecies except Meitanaphis microgallis using 20 gene regions: five nuclear genes as well as 13 protein-coding genes and two rRNA genes …

Student-Faculty Collaborative Research Grant Report, Megan Bestwick

Post-Grant Reports

Mitochondria are essential organelles in most eukaryotic cells because of their role in metabolism and the production of ATP by the oxidative phosphorylation (OXPHOS) pathway, as well as other key cellular processes. Metal cofactors, such as copper (Cu) and iron (Fe), are incorporated into OXPHOS protein complexes of yeast located within the inner membrane of the mitochondria. Misincorporation or modulation of these available metals in mitochondrial enzymes leads to the production of reactive oxygen species (ROS). ROS are reactive molecules containing oxygen such as peroxides, superoxide, and hydroxyl radicals. Yeast are a good model for studying aging and the effect …

Identification Of The Alternative Oxidase Gene And Its Expression In The Copepod Tigriopus Californicus, Allison Mcdonald, Carly E. Tward, Willie Cygelfarb, Jaspreet Singh

Biology Faculty Publications

In addition to the typical electron transport system (ETS) in animal mitochondria responsible for oxidative phosphorylation, in some species there exists an alternative oxidase (AOX) pathway capable of catalyzing the oxidation of ubiquinol and the reduction of oxygen to water. The discovery of AOX in animals is recent and further investigations into its expression, regulation, and physiological role have been hampered by the lack of a tractable experimental model organism. Our recent DNA database searches using bioinformatics revealed an AOX sequence in several marine copepods including Tigriopus californicus. This species lives in tidepools along the west coast of North …

Effects Of Commercial Formulations Of Glyphosate On Saccharomyces Cerevisiae, Apoorva Ravi Shankar

Graduate Theses, Dissertations, and Problem Reports (ETD)

Commercial formulations of glyphosate are among the most extensively used herbicides around the world. The active ingredient, glyphosate, targets the aromatic amino acid pathway. This pathway is absent in mammals, resulting in low toxicity. Different formulations contain varying adjuvants and surfactants, whose synergistic effects are yet to be extensively studied at the cellular level. In this study, I tested multiple commercial formulations that showed a variation in growth phenotype among different yeast strains. To gain a better understanding of response and resistance mechanisms at the genome and transcriptome level, I carried out an in-lab evolution study, along with a transcriptome …

Perturbation Of Energy Metabolism At The Center Of The Mechanism Of Action Of Valproate, Michael Ghassan Salsaa

Wayne State University Dissertations

Bipolar disorder (BD) is a common and debilitating psychiatric disorder. Valproic acid (VPA) is one of the major drugs used to treat BD patients. However, it is not universally effective and, in addition, causes severe side effects. Its mechanism of action is not known, which complicates efforts to develop more effective drugs. Studies have established that VPA perturbs metabolism, which is implicated in both the therapeutic mechanism of action of the drug as well as drug toxicity. However, the mechanism whereby VPA causes these perturbations is not understood. To address this knowledge gap, I investigated the acute and chronic effects …

Genetic Basis Of Thermal Divergence In Saccharomyces Species, Xueying C. Li

Arts & Sciences Graduate Student Theses and Dissertations

The genetic architecture of phenotypic divergence is a central question in evolutionary biology. Genetic architecture is impacted by whether evolution occurs through accumulation of many small-effect or a few large-effect changes, the relative contribution of coding and cis-regulatory changes, and the prevalence of epistatic effects. Our empirical understanding of the genetic basis of evolutionary change remains incomplete, largely because reproductive barriers limit genetic analysis to those phenotypes that distinguish closely related species. In this dissertation, I use hybrid genetic analysis to examine the basis of thermal divergence between two post-zygotically isolated species, Saccharomyces cerevisiae and S. uvarum. S. cerevisiae is …

Mechanisms Of S1p-Induced Endothelial Barrier Enhancement, Natascha Guimarães Alves

USF Tampa Graduate Theses and Dissertations

Excessive microvascular permeability is a serious complication involved in traumatic injury and inflammatory diseases. Alcohol intoxication can exacerbate the physiological derangements produced by microvascular endothelial barrier dysfunction in such disease conditions. Sphingosine-1-phosphate (S1P) has known endothelial barrier-protective properties, and has been shown to ameliorate microvascular leakage in a model of combined alcohol intoxication and hemorrhagic shock and resuscitation (HSR). However, whether the barrier-protective properties of S1P extend to endothelial cells of the blood-brain barrier (BBB) is unclear. The mechanisms of S1P-induced barrier protection during alcohol intoxication or HSR are also unknown. In the current study, we tested the hypothesis that …

Anti-Apoptotic Effects Of Human Placental Hydrolysate Against Hepatocyte Toxicity In Vivo And In Vitro, Dong Ho Bak, Jungtae Na, Mi Ji Choi, Byung Chul Lee, Chang Taek Oh, Jeom Yong Kim, Hae Jung Han, Moo Joong Kim, Tae Ho Kim, Beom Joon Kim

Biological Sciences Faculty Publications

Apoptosis and oxidative stress are essential for the pathogenesis of acute liver failure and fulminant hepatic failure. Human placental hydrolysate (hPH) has been reported to possess antioxidant and anti-inflammatory properties. In the present study, the protective effects of hPH against D-galactosamine (D-GalN)- and lipopolysaccharide (LPS)-induced hepatocyte apoptosis were investigated in vivo. In addition, the molecular mechanisms underlying the anti-apoptotic activities of hPH against D-GalN-induced cell death in vitro were examined. Male Sprague-D awley rats were injected with D-GaIN/LPS with or without the administration of hPH. Rats were sacrificed 24 h after D-GaIN/LPS intraperitoneal injection, and the blood and liver samples …

Evidence That A Respiratory Shield In Escherichia Coli Protects A Low-Molecular-Mass Fe-Ii Pool From O2-Dependent Oxidation, Joshua D. Wofford, Naimah Bolaji, Nathaniel Dziuba, Franklin Wayne Outten, Paul A. Lindahl

Faculty Publications

Iron is critical for virtually all organisms, yet major questions remain regarding the systems' level understanding of iron in whole cells. Here, we obtained Mössbauer and EPR spectra of Escherichia coli cells prepared under different nutrient iron concentrations, carbon sources, growth phases, and O₂ concentrations to better understand their global iron content. We investigated wild-type cells and those lacking Fur, FtnA, Bfr, and Dps proteins. The coarse-grain iron content of exponentially growing cells consisted of iron-sulfur clusters, variable amounts of nonheme high-spin Fe^II species, and an unassigned residual quadrupole doublet. The iron in stationary-phase cells was dominated by …

Importance Of S. Cerevisiae Rcf1 And Rcf2 Proteins For The Mitochondrial Protonmotive Force Generation, Vera Strogolova

Dissertations (1934 -)

Mitochondria are the site of oxidative phosphorylation (OXPHOS) pathway, which can supply majority of energy in a eukaryotic cell. OXPHOS enzyme activities generate electrochemical gradient known as mitochondrial protonmotive force (PMF). PMF coordinates OXPHOS enzyme activities and supports essential cell survival functions such as transport of proteins and metabolites in and out of mitochondria. PMF is maintained despite variations in cellular energy demand and oxygen availability.Mitochondrial proteins belonging to the conserved hypoxia induced gene domain (HIGD) family improve cell survival during the hypoxic and hypoglycemic stress. Their molecular function is not fully understood but they seem to act through …

A Disturbance In The Force: Cellular Stress Sensing By The Mitochondrial Network, Robert Gilkerson

School of Integrative Biological & Chemical Sciences (Formerly Dept. of Biology)

As a highly dynamic organellar network, mitochondria are maintained as an organellar network by delicately balancing fission and fusion pathways. This homeostatic balance of organellar dynamics is increasingly revealed to play an integral role in sensing cellular stress stimuli. Mitochondrial fission/fusion balance is highly sensitive to perturbations such as loss of bioenergetic function, oxidative stress, and other stimuli, with mechanistic contribution to subsequent cell-wide cascades including inflammation, autophagy, and apoptosis. The overlapping activity with m-AAA protease 1 (OMA1) metallopeptidase, a stress-sensitive modulator of mitochondrial fusion, and dynamin-related protein 1 (DRP1), a regulator of mitochondrial fission, are key factors that shape …

Functional Variation In The Mitochondrial Genome Of The Yeast Saccharomyces Cerevisiae, John F. Wolters

Graduate Dissertations and Theses

Mitochondrial haplotypes contribute to functional diversity in natural populations. Uniparental inheritance makes it difficult to characterize the genetic architecture of mitochondrially driven phenotypes. In this work I explored the natural diversity of mitochondrial genomes in the yeast Saccharomyces cerevisiae. Few complete mitochondrial genomes were available for S. cerevisiae owing to challenges in high-throughput sequencing. I developed sequencing strategies using new technologies to generate complete high quality yeast mitochondrial genomes. Comparisons of 100 complete yeast mitochondrial genomes demonstrated extensive variation between populations in coding sequences and variable introns.

I demonstrated that these mitochondrial variants directly caused growth differences in strains with …

Mitochondrial Dynamics And Respiration Within Cells With Increased Open Pore Cytoskeletal Meshes, David H. Jang, Sarah C. Seeger, Martha E. Grady, Frances S. Shofer, David M. Eckmann

Mechanical Engineering Faculty Publications

The cytoskeletal architecture directly affects the morphology, motility, and tensional homeostasis of the cell. In addition, the cytoskeleton is important for mitosis, intracellular traffic, organelle motility, and even cellular respiration. The organelle responsible for a majority of the energy conversion for the cell, the mitochondrion, has a dependence on the cytoskeleton for mobility and function. In previous studies, we established that cytoskeletal inhibitors altered the movement of the mitochondria, their morphology, and their respiration in human dermal fibroblasts. Here, we use this protocol to investigate applicability of power law diffusion to describe mitochondrial locomotion, assessment of rates of fission and …

The Landscape Of Mtdna Modifications In Cancer: A Tale Of Two Cities, Kate L. Hertweck, Santanu Dasgupta

Biology Faculty Publications and Presentations

Mitochondria from normal and cancerous cells represent a tale of two cities, wherein both execute similar processes but with different cellular and molecular effects. Given the number of reviews currently available which describe the functional implications of mitochondrial mutations in cancer, this article focuses on documenting current knowledge in the abundance and distribution of somatic mitochondrial mutations, followed by elucidation of processes which affect the fate of mutations in cancer cells. The conclusion includes an overview of translational implications for mtDNA mutations, as well as recommendations for future research uniting mitochondrial variants and tumorigenesis.

Oxidative Insults Disrupt Mitochondrial Dynamics, Iraselia A. Garcia

Theses and Dissertations

Mitochondria form an organellar network to provide ATP to the cell. Mitochondrial DNA (mtDNA) combines with nuclear DNA to encode polypeptides critical to forming the complexes of oxidative phosphorylation in the mitochondrial inner membrane, which generate a transmembrane potential (ΔΨm) to synthesize ATP. This ΔΨm is required to maintain mitochondria fission/fusion dynamics: organellar fusion (mediated by OPA1) and fission events (mediated by DRP1) coordinately regulate mitochondrial dynamics. While oxidative stress correlates with mitochondrial dysfunction, it is unclear how oxidants affect mitochondrial structure/function homeostasis. This project seeks to establish and examine the impact of reactive oxygen species (ROS) on mitochondrial dynamics. …

Mitochondrial Dynamics Controls T Cell Fate Through Metabolic Programming, Michael Buck

Arts & Sciences Graduate Student Theses and Dissertations

Activated effector T (TE) cells augment anabolic pathways of metabolism, such as aerobic glycolysis, while memory T (TM) cells engage catabolic pathways, like fatty acid oxidation (FAO). However, signals that drive these differences remain unclear. Mitochondria are metabolic organelles that actively transform their ultrastructure. Therefore, we questioned whether mitochondrial dynamics controls T cell metabolism. We show that TE cells have punctate mitochondria, while TM cells maintain fused networks. The fusion protein Opa1 is required for TM, but not TE cells after infection, and enforcing fusion in TE cells imposes TM cell characteristics and enhances antitumor function. Our data suggest that, …

Mitochondrial Damage Accumulation In Oocytes – A Potential Link Between Maternal Obesity And Increased Cardiometabolic Disease Risk In Offspring., Anna Louise Boudoures

Arts & Sciences Graduate Student Theses and Dissertations

The developmental origins of health and disease (DoHAD) hypothesis suggests that negative maternal lifestyle choices, such as obesity, affect the health of her offspring. Clinical and laboratory studies support this hypothesis – offspring born to obese mothers are at increased risk for health conditions including cardiometabolic syndrome and congenital abnormalities. Maternal obesity damages the oocytes, contributing to the increased disease risk by transmitting damaged organelles and epigenetic modifications to the offspring. Mitochondria, the most abundant organelle in the oocyte, are damaged in oocytes from obese females. However, we do not understand if mitochondrial damage in oocytes is reversible nor why …

Mutational Analysis Of The Qrrq Motif In The Yeast Hig1 Type 2 Protein Rcf1 Reveals A Regulatory Role For The Cytochrome C Oxidase Complex, Joshua Garlich, Valentina Strecker, Ilka Wittig, Rosemary A. Stuart

Biological Sciences Faculty Research and Publications

The yeast Rcf1 protein is a member of the conserved family of proteins termed the hypoxia-induced gene (domain) 1 (Hig1 or HIGD1) family. Rcf1 interacts with components of the mitochondrial oxidative phosphorylation system, in particular the cytochrome bc₁(complex III)-cytochrome c oxidase (complex IV) supercomplex (termed III-IV) and the ADP/ATP carrier proteins. Rcf1 plays a role in the assembly and modulation of the activity of complex IV; however, the molecular basis for how Rcf1 influences the activity of complex IV is currently unknown. Hig1 type 2 isoforms, which include the Rcf1 protein, are characterized in part by the presence …

Reduced Mitochondrial Efficiency Explains Mismatched Growth And Metabolic Rate At Supraoptimal Temperatures., Eloy Martinez, Michael A. Menze, Salvatore J. Agosta

Faculty and Staff Scholarship

The relationship between whole-organism growth and metabolism is generally assumed to be positive and causative; higher metabolic rates support higher growth rates. In Manduca sexta, existing data demonstrate a deviation from this simple prediction: at supraoptimal temperatures for larval growth, metabolic rate keeps increasing while growth rate is decreasing. This mismatch presumably reflects the rising “cost of maintenance” with temperature. Precisely what constitutes this cost is not clear, but we suspect the efficiency with which mitochondria harness oxygen and organic substrates into cellular energy (ATP) is key. We tested this by integrating existing data on M. sexta growth and …

Effect Of Trehalose As An Additive To Dimethyl Sulfoxide Solutions On Ice Formation, Cellular Viability, And Metabolism., Jason Solocinski, Quinn Osgood, Mian Wang, Aaron Connolly, Michael A. Menze, Nilay Chakraborty

Faculty and Staff Scholarship

Cryopreservation is the only established method for long-term preservation of cells and cellular material. This technique involves preservation of cells and cellular components in the presence of cryoprotective agents (CPAs) at liquid nitrogen temperatures (−196 °C). The organic solvent dimethyl sulfoxide (Me₂SO) is one of the most commonly utilized CPAs and has been used with various levels of success depending on the type of cells. In recent years, to improve cryogenic outcomes, the non-reducing disaccharide trehalose has been used as an additive to Me₂SO-based freezing solutions. Trehalose is a naturally occurring non-toxic compound found in bacteria, …

Msl1 Is A Mechanosensitive Ion Channel That Dissipates Mitochondrial Membrane Potential And Maintains Redox Homeostasis In Mitochondria During Abiotic Stress, Chun Pong Lee, Grigory Maksaev, Gregory S. Jensen, Monika W. Murcha, Margaret E. Wilson, Mark Fricker, Ruediger Hell, Elizabeth S. Haswell, A Harvey Millar, Lee J. Sweetlove

Biology Faculty Research

Mitochondria must maintain tight control over the electrochemical gradient across their inner membrane to allow ATP synthesis while maintaining a redox-balanced electron transport chain and avoiding excessive reactive oxygen species production. However, there is a scarcity of knowledge about the ion transporters in the inner mitochondrial membrane that contribute to control of membrane potential. We show that loss of MSL1, a member of a family of mechanosensitive ion channels related to the bacterial channel MscS, leads to increased membrane potential of Arabidopsis mitochondria under specific bioenergetic states. We demonstrate that MSL1 localises to the inner mitochondrial membrane. When expressed in …

Subcellular And In-Vivo Nano-Endoscopy, Surya Venkatasekhar Cheemalapati, John Winskas, Hao Wang, Karthik Konnaiyan, Arseny Zhdanov, Alison Roth, Swamy Rakesh Adapa, Andrew Deonarine, Rays H. Y. Jiang, Anna Pyayt

Chemical, Biological and Materials Engineering Faculty Publications

Analysis of individual cells at the subcellular level is important for understanding diseases and accelerating drug discovery. Nanoscale endoscopes allow minimally invasive probing of individual cell interiors. Several such instruments have been presented previously, but they are either too complex to fabricate or require sophisticated external detectors because of low signal collection efficiency. Here we present a nanoendoscope that can locally excite fluorescence in labelled cell organelles and collect the emitted signal for spectral analysis. Finite Difference Time Domain (FDTD) simulations have shown that with an optimized nanoendoscope taper profile, the light emission and collection was localized within ~100 nm. …

Characterization Of The Rcf1 Q-X3-R-X-R-X3-Q Motif And Its Role In Supporting Oxidative Phosphorylation Enzyme Complexes, Joshua Garlich

Dissertations (1934 -)

Mitochondrial respiratory complexes are critical components of cellular energy production that require tight regulation to ensure optimal function. Rcf1 and Rcf2 are mitochondrial proteins that can physically associate with the yeast respiratory complexes III and IV, and the higher-ordered III-IV respiratory supercomplex that also contains the ADP/ATP translocase, AAC. Rcf1 can physically associate with both complex III and IV independently, and can be chemically crosslinked to AAC, indicating a close physical proximity to a predominant regulator of energy flux within the cell. It was therefore hypothesized that Rcf1, through its physical association with complexes III and IV, and its close …

Neofunctionalization Of Dual-Targeted Plant Organelle Dna Replication Proteins

Journal of Undergraduate Research

Neofunctionalization is the process by which genes gain a new or modified function as a result of gene duplication and modification of one or more of the new genes. This exciting area of research helps explain the diversification of protein functions in cells. The number of documented examples has increased in recent years in all classes of organisms (Deng et al., 2010; Han et al. 2011; Cannela et al. 2014; Sakuma et al. 2013; Zhang et al. 2014). In some cases a protein that was previously thought to function in only one cellular compartment has been shown to also function …

Mir494 Reduces Renal Cancer Cell Survival Coinciding With Increased Lipid Droplets And Mitochondrial Changes, Punashi Dutta, Edward Haller, Arielle Sharp, Meera Nanjundan

Molecular Biosciences Faculty Publications

Background: miRNAs can regulate cellular survival in various cancer cell types. Recent evidence implicates the formation of lipid droplets as a hallmark event during apoptotic cell death response. It is presently unknown whether MIR494, located at 14q32 which is deleted in renal cancers, reduces cell survival in renal cancer cells and if this process is accompanied by changes in the number of lipid droplets.

Methods: 769-P renal carcinoma cells were utilized for this study. Control or MIR494 mimic was expressed in these cells following which cell viability (via crystal violet) and apoptotic cell numbers (via Annexin V/PI staining) were …

Sodium Borate Treatment Induces Metabolic Reprogramming In Hepatocellular Carcinoma Through Sirt3 Activation, Berna Üstüner, Hüseyi̇n Çi̇men

Turkish Journal of Biology

Sirtuins are NAD+-dependent deacetylases and ADP ribosyltransferases that are activated under stress conditions such as calorie restriction and starvation. Boron, which is mostly found in the form of boric acid (BA) or sodium borate (NaB), is known to bind NAD+. In this study, the effect of NaB on hepatoma cell line HEP3B was investigated by analyzing the proteins harvested from NaB-treated and serum-starved HEP3B cells. The NaB treatment (15 μg/mL) led to a decrease in the overall proteome acetylation and particularly in mitochondria, the synthesis rate of respiratory complexes, the amount of cellular reactive oxygen species, and the proliferation rate …

Functional Significance Of Mtdna Cytosine Modification Tested By Genome Editing, Jason M. Robinson

Theses and Dissertations

The field of epigenetics is gaining popularity and speed, due in part to its capability to answer lingering questions about the root cause of certain diseases. Epigenetics plays a crucial role in regulation of the cell and cell survival, particularly by cytosine methylation. It remains controversial if DNMT’s which facilitate methylation are present in mammalian mitochondria and what the functional significance they may have on modification of mitochondrial DNA. CRISPR-Cas9 technology enabled genome editing to remove the MTS (mitochondrial targeting sequence) from DNMT1 of HCT116 cells, purposefully minimizing effects on nuclear cytosine methylation, while exclusively impacting mitochondrial modification. Removal of …

Two Complete Mitochondrial Genomes From Praticolella Mexicana Perez, 2011 (Polygyridae) And Gene Order Evolution In Helicoidea (Mollusca, Gastropoda), Russell L. Minton, Marco A. Martinez Cruz, Mark L. Farman, Kathryn E. Perez

School of Integrative Biological & Chemical Sciences (Formerly Dept. of Biology)

Helicoidea is a diverse group of land snails with a global distribution. While much is known regarding the relationships of helicoid taxa, comparatively little is known about the evolution of the mitochondrial genome in the superfamily. We sequenced two complete mitochondrial genomes from Praticolella mexicana Perez, 2011 representing the first such data from the helicoid family Polygyridae, and used them in an evolutionary analysis of mitogenomic gene order. We found the mitochondrial genome of P. mexicana to be 14,008 bp in size, possessing the typical 37 metazoan genes. Multiple alternate stop codons are used, as are incomplete stop codons. Mitogenome …