scholarly journals Involvement of a G Protein Regulatory Circuit in Alternative Oxidase Production in Neurospora crassa

2019 ◽  
Vol 9 (10) ◽  
pp. 3453-3465 ◽  
Author(s):  
Natasa Bosnjak ◽  
Kristina M. Smith ◽  
Iman Asaria ◽  
Adrian Lahola-Chomiak ◽  
Nishka Kishore ◽  
...  
Keyword(s):  
Electron Transport ◽  
Neurospora Crassa ◽  
G Protein ◽  
Translational Control ◽  
Electron Transport Chain ◽  
Alternative Oxidase ◽  
G Protein Signaling ◽  
Mrna Levels ◽  
Protein Signaling ◽  
Transport Chain

The Neurospora crassa nuclear aod-1 gene encodes an alternative oxidase that functions in mitochondria. The enzyme provides a branch from the standard electron transport chain by transferring electrons directly from ubiquinol to oxygen. In standard laboratory strains, aod-1 is transcribed at very low levels under normal growth conditions. However, if the standard electron transport chain is disrupted, aod-1 mRNA expression is induced and the AOD1 protein is produced. We previously identified a strain of N. crassa, that produces high levels of aod-1 transcript under non-inducing conditions. Here we have crossed this strain to a standard lab strain and determined the genomic sequences of the parents and several progeny. Analysis of the sequence data and the levels of aod-1 mRNA in uninduced cultures revealed that a frameshift mutation in the flbA gene results in the high uninduced expression of aod-1. The flbA gene encodes a regulator of G protein signaling that decreases the activity of the Gα subunit of heterotrimeric G proteins. Our data suggest that strains with a functional flbA gene prevent uninduced expression of aod-1 by inactivating a G protein signaling pathway, and that this pathway is activated in cells grown under conditions that induce aod-1. Induced cells with a deletion of the gene encoding the Gα protein still have a partial increase in aod-1 mRNA levels, suggesting a second pathway for inducing transcription of the gene in N. crassa. We also present evidence that a translational control mechanism prevents production of AOD1 protein in uninduced cultures.

Modulation of renal tubular cell function by RGS3

1999 ◽  
Vol 276 (4) ◽  
pp. F535-F543 ◽  
Author(s):  
W. Grüning ◽  
T. Arnould ◽  
F. Jochimsen ◽  
L. Sellin ◽  
S. Ananth ◽  
...  
Keyword(s):  
G Protein ◽  
Cell Function ◽  
Collecting Duct ◽  
Inducible Promoter ◽  
Polyclonal Antiserum ◽  
Renal Tubular Cell ◽  
G Protein Signaling ◽  
Mrna Levels ◽  
Protein Signaling ◽  
Adult Kidney

The recently discovered family of regulators of G protein signaling (RGS) accelerates the intrinsic GTPase activity of certain Gα subunits, thereby terminating G protein signaling. Particularly high mRNA levels of one family member, RGS3, are found in the adult kidney. To establish the temporal and spatial renal expression pattern of RGS3, a polyclonal antiserum was raised against the COOH terminus of RGS3. Staining of mouse renal tissue at different gestational stages revealed high levels of RGS3 within the developing and mature tubular epithelial cells. We tested whether RGS3 can modulate tubular migration, an important aspect of tubular development, in response to G protein-mediated signaling. Several mouse intermedullary collecting duct (mIMCD-3) cell lines were generated that expressed RGS3 under the control of an inducible promoter. Lysophosphatidic acid (LPA) is a potent chemoattractant that mediates its effects through heterotrimeric G proteins. We found that induction of RGS3 significantly reduced LPA-mediated cell migration in RGS3-expressing mIMCD-3 clones, whereas chemotaxis induced by hepatocyte growth factor remained unaffected by RGS3. Our findings suggest that RGS3 modulates tubular functions during renal development and in the adult kidney.

scholarly journals Mitochondrial miR-181a-5p promotes glucose metabolism reprogramming in liver cancer by regulating the electron transport chain

2019 ◽  
Vol 41 (7) ◽  
pp. 972-983 ◽  
Author(s):  
Xiang Zhuang ◽  
Yuwei Chen ◽  
Zhenru Wu ◽  
Qing Xu ◽  
Menglin Chen ◽  
...  
Keyword(s):  
Electron Transport ◽  
Glucose Metabolism ◽  
Liver Cancer ◽  
Electron Transport Chain ◽  
Glucose Transporter ◽  
Mitochondrial Gene ◽  
Mrna Levels ◽  
Atp Production ◽  
Transport Chain

Abstract Liver cancer and other malignant tumor cells rely on the glycolytic pathway to obtain energy (i.e. the Warburg effect); however, the underlying mechanism is unclear. Mitochondria are sites of oxidative phosphorylation and adenosine triphosphate (ATP) production. The 13 constituent respiratory chain proteins encoded by the mitochondrial genome (namely, mtDNA) play essential roles. We found that in human hepatocellular carcinoma (HCC) tissues, 11 out of the 13 mtDNA-encoded genes exhibited decreased mRNA levels and 5 genes displayed decreased protein levels, including the cytochrome B (mt-CYB) and cytochrome C oxidase II (mt-CO2) genes. Mitochondrial gene sequencing revealed abnormalities in the levels of a large number of mitochondrial miRNAs (mitomiRs). MicroRNA-181a-5p (mir-181a-5p), which potentially targets genes encoding mt-CYB and mt-CO2 protein, was screened out from 549 downregulated mitomiRs via bioinformatic analysis. After overexpression of mitomiR-181a-5p, mt-CYB and mt-CO2 levels were reduced in HCC cells, and the mitochondrial membrane potential (MMP) maintained by the electron transport chain (ETC) was decreased. Furthermore, the expression of hexokinase 2 (HK2) and glucose transporter type 1 (GLUT1) was upregulated, accompanied by elevated glucose, lactic acid release, and activity of lactate dehydrogenase (LDH). In vivo experiments confirmed that constitutive mitomiR-181a-5p expression caused reprogramming of glucose metabolism and promoted tumor growth and early lung metastasis in liver cancer. In summary, the present study reveals the important role of mitomiRs in glucose metabolism reprogramming in liver cancer, which is of considerable value in exploring new therapeutic targets for HCC.

Abstract P177: Reduced Expression of Regulator of G-Protein Signaling-2 (RGS2) in the Placenta During Preeclampsia

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Katherine J Perschbacher ◽  
Donna A Santillan ◽  
Eric J Devor ◽  
Sabrina M Scroggins ◽  
Jeremy A Sandgren ◽  
...  
Keyword(s):  
G Protein ◽  
Association Studies ◽  
Tissue Bank ◽  
Placental Tissue ◽  
G Protein Signaling ◽  
Wildtype Mouse ◽  
Rgs Proteins ◽  
Mrna Levels ◽  
Protein Signaling ◽  
Increased Risk

Preeclampsia (PE) is a serious cardiovascular condition of late pregnancy. Genetic risk factors and the early-gestational etiology remain largely unclear, though evidence supports excessive activation of Gαq signaling within the placenta in response to various hormones including vasopressin, endothelin, and angiotensin. Regulator of G-protein Signaling 2 (RGS2) acts as an endogenous terminator of Gαq signaling, and previous association studies have identified an increased risk for PE and its sequelae in women carrying a single nucleotide polymorphism that is expected to reduce levels of RGS2. We hypothesized that RGS2 is expressed in placental trophoblasts, and that reduced expression of RGS2 in placental tissue may represent a risk factor for the development of PE. Whole placenta samples and clinical data from preeclamptic and clinically-matched control pregnancies were obtained from the University of Iowa Maternal-Fetal Tissue Bank (IRB#200910784) and examined for mRNA levels of the B/R4 family of RGS proteins, including RGS2. Of the members examined (RGS2, -3, -4.2, -4.3, -4.4, -4.5, and -5) in control placentas (n=9), only RGS2 (Ct 28.8±0.7 vs 18S Ct 12.1±0.4) and RGS4.3 (Ct 23.0±0.4 vs 18S Ct 13.3±0.3) transcripts were expressed above background levels. RGS2 protein expression was then confirmed in human placental tissues by Western blot. RGS2 mRNA expression was 3-fold higher in fetal (amniotic, p<0.05) layers than maternal (decidual) layers. In preeclamptic placenta (n=11), RGS2 may be suppressed (1.0±0.4 vs 0.2±0.3-fold, p=0.1) while RGS4.3 remains unchanged (1.0±0.4 vs 1.1±0.4 fold, p=0.8). Initial immunohistochemical detection confirms cytoplasmic localization of RGS2 in trophoblasts of wildtype mouse placenta, despite exclusive nuclear localization in other tissues. We conclude that human placenta expresses RGS2, and that this expression may be suppressed during preeclampsia. Loss of RGS2 expression may result in disinhibited trophoblast Gαq signaling, and ultimately placental insufficiency.

Metabolic Acidosis Regulates RGS16 and G-protein Signaling in Osteoblasts

2021 ◽  
Author(s):  
Nancy S. Krieger ◽  
David A. Bushinsky
Keyword(s):  
Metabolic Acidosis ◽  
Bone Resorption ◽  
G Protein ◽  
Specific Inhibitor ◽  
Osteoclastic Bone Resorption ◽  
G Protein Signaling ◽  
Rgs Proteins ◽  
Mrna Levels ◽  
Protein Signaling ◽  
Stimulation Of

Chronic metabolic acidosis stimulates cell-mediated net calcium efflux from bone mediated by increased osteoblastic cyclooxygenase 2 (COX2), leading to prostaglandin E2-induced stimulation of RANKL-induced osteoclastic bone resorption. The osteoblastic H+-sensing G-protein coupled receptor (GPCR), OGR1, is activated by acidosis and leads to increased bne resorption. As regulators of G protein signaling (RGS) proteins limit GPCR signaling, we tested whether RGS proteins themselves are regulated by metabolic acidosis. Primary osteoblasts were isolated from neonatal mouse calvariae and incubated in physiological neutral (NTL) or acidic (MET) medium. Cells were collected and RNA extracted for real time PCR analysis with mRNA levels normalized to RPL13a. RGS1, RGS2, RGS3, RGS4, RGS10, RGS11 or RGS18mRNA did not differ between MET and NTL; however by 30' MET decreased RGS16 which persisted for 60' and 3h. Incubation of osteoblasts with the OGR1 inhibitor CuCl2 inhibited the MET induced increase in RGS16 mRNA. Gallein, a specific inhibitor of Gβγ signaling, was used to determine if downstream signaling by the βγ subunit was critical for the response to acidosis. Gallein decreased net Ca efflux from calvariae and COX2 and RANKL gene expression from isolated osteoblasts. These results indicate that regulation of RGS16 plays an important role in modulating the response of the osteoblastic GPCR, OGR1, to metabolic acidosis and subsequent stimulation of osteoclastic bone resorption.

scholarly journals RgsA Attenuates the PKA Signaling, Stress Response, and Virulence in the Human Opportunistic Pathogen Aspergillus fumigatus

2019 ◽  
Vol 20 (22) ◽  
pp. 5628 ◽  
Author(s):  
Hnin Phyu Lwin ◽  
Yong-Ho Choi ◽  
Min-Woo Lee ◽  
Jae-Hyuk Yu ◽  
Kwang-Soo Shin
Keyword(s):  
Stress Response ◽  
Aspergillus Fumigatus ◽  
G Protein ◽  
Stress Responses ◽  
Pathogenic Fungus ◽  
G Protein Signaling ◽  
Mrna Levels ◽  
Protein Signaling ◽  
Null Mutant ◽  
Genes Encoding

The regulator of G-protein signaling (RGS) proteins play an important role in upstream control of heterotrimeric G-protein signaling pathways. In the genome of the human opportunistic pathogenic fungus Aspergillus fumigatus, six RGS protein-encoding genes are present. To characterize the rgsA gene predicted to encode a protein with an RGS domain, we generated an rgsA null mutant and observed the phenotypes of the mutant. The deletion (Δ) of rgsA resulted in increased radial growth and enhanced asexual sporulation in both solid and liquid culture conditions. Accordingly, transcripts levels of the key asexual developmental regulators abaA, brlA, and wetA are elevated in the ΔrgsA mutant. Moreover, ΔrgsA resulted in elevated spore germination rates in the absence of a carbon source. The activity of cAMP-dependent protein kinase A (PKA) and mRNA levels of genes encoding PKA signaling elements are elevated by ΔrgsA. In addition, mRNA levels of genes associated with stress-response signaling increased with the lack of rgsA, and the ΔrgsA spores showed enhanced tolerance against oxidative stressors. Comparative transcriptomic analyses revealed that the ΔrgsA mutant showed higher mRNA levels of gliotoxin (GT) biosynthetic genes. Accordingly, the rgsA null mutant exhibited increased production of GT and elevated virulence in the mouse. Conversely, the majority of genes encoding glucan degrading enzymes were down-regulated by ΔrgsA, and endoglucanase activities were reduced. In summary, RgsA plays multiple roles, governing growth, development, stress responses, virulence, and external polymer degradation—likely by attenuating PKA signaling.

scholarly journals Mitochondrial Electron Transport Chain Inhibition Suppresses LPS-Induced Inflammatory Responses via TREM1/STAT3 Pathway in BV2 Microglia

2019 ◽  
Author(s):  
Cuiyan Zhou ◽  
Jie Zhang ◽  
Weihai Ying
Keyword(s):  
Electron Transport ◽  
Electron Transport Chain ◽  
Microglial Activation ◽  
Nuclear Translocation ◽  
Inflammatory Responses ◽  
Mitochondrial Damage ◽  
Mrna Levels ◽  
Mitochondrial Electron Transport Chain ◽  
Transport Chain ◽  
Bv2 Microglia

AbstractMitochondrial damage and neuroinflammation belong to two of the most important pathological factors in multiple neurological disorders. However, the effect of mitochondrial damage of microglia on microglial activation under pathological conditions has remained unclear. In our current study, we used BV2 microglia as a cellular model to determine the effects of mitochondrial electron transport chain (ETC) inhibitors on LPS-induced inflammatory responses of microglia. We found that all of the three mitochondrial ETC inhibitors, including rotenone, sodium azide and antimycin A, significantly inhibited LPS-induced inflammatory responses of the microglia, assessed by determinations of the protein or mRNA levels of IL-1β, IL-6, TNF-α, iNOS and COX2. Nuclear translocation of NF-κB p65 subunit does not appear to play an important role in the mitochondrial ETC inhibition-produced suppression of microglial activation. Instead, our study found that the mitochondrial ETC inhibitors significantly attenuated not only the LPS-induced increase in the TREM1 levels - an amplifier of inflammatory process, but also the LPS-induced increase in the ratio of phosphorylated STAT3 / STAT3. In summary, our study has suggested that mitochondrial ETC inhibition of microglia can lead to suppression of LPS-induced microglial activation, which may be mediated by the inhibitory effects of mitochondrial ETC inhibition on the LPS-induced increases in the level of TREM1 and the ratio of p-STAT3 / STAT3. These findings have provided valuable information for elucidating the relationships between mitochondrial damage and neuroinflammation in multiple neurological diseases.

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