scholarly journals Single cell functional genomics reveals the importance of mitochondria in cell-to-cell variation in proliferation, drug resistance and mutation outcome

2018 ◽  
Author(s):  
Riddhiman Dhar ◽  
Alsu M Missarova ◽  
Ben Lehner ◽  
Lucas B Carey
Keyword(s):  
Drug Resistance ◽  
Single Cell ◽  
Stress Tolerance ◽  
Mammalian Cells ◽  
Wild Type ◽  
Common Environment ◽  
Automated Microscopy ◽  
Cell Variation ◽  
Anti Fungal ◽  
The Impact

AbstractMutations frequently have outcomes that differ across individuals, even when these individuals are genetically identical and share a common environment. Moreover, individual microbial and mammalian cells can vary substantially in their proliferation rates, stress tolerance, and drug resistance, with important implications for the treatment of infections and cancer. To investigate the causes of cell-to-cell variation in proliferation, we developed a high-throughput automated microscopy assay and used it to quantify the impact of deleting >1,500 genes in yeast. Mutations affecting mitochondria were particularly variable in their outcome. In both mutant and wild-type cells mitochondria state – but not content – varied substantially across individual cells and predicted cell-to-cell variation in proliferation, mutation outcome, stress tolerance, and resistance to a clinically used anti-fungal drug. These results suggest an important role for cell-to-cell variation in the state of an organelle in single cell phenotypic variation.

scholarly journals Single cell functional genomics reveals the importance of mitochondria in cell-to-cell phenotypic variation

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Riddhiman Dhar ◽  
Alsu M Missarova ◽  
Ben Lehner ◽  
Lucas B Carey
Keyword(s):  
Single Cell ◽  
Stress Tolerance ◽  
Phenotypic Variation ◽  
Mammalian Cells ◽  
Wild Type ◽  
Common Environment ◽  
Automated Microscopy ◽  
Cell Variation ◽  
Anti Fungal ◽  
The Impact

Mutations frequently have outcomes that differ across individuals, even when these individuals are genetically identical and share a common environment. Moreover, individual microbial and mammalian cells can vary substantially in their proliferation rates, stress tolerance, and drug resistance, with important implications for the treatment of infections and cancer. To investigate the causes of cell-to-cell variation in proliferation, we used a high-throughput automated microscopy assay to quantify the impact of deleting >1500 genes in yeast. Mutations affecting mitochondria were particularly variable in their outcome. In both mutant and wild-type cells mitochondrial membrane potential – but not amount – varied substantially across individual cells and predicted cell-to-cell variation in proliferation, mutation outcome, stress tolerance, and resistance to a clinically used anti-fungal drug. These results suggest an important role for cell-to-cell variation in the state of an organelle in single cell phenotypic variation.

scholarly journals Single-cell replication profiling reveals stochastic regulation of the mammalian replication-timing program

2017 ◽  
Author(s):  
Vishnu Dileep ◽  
David M. Gilbert
Keyword(s):  
Single Cell ◽  
Mammalian Cells ◽  
Replication Timing ◽  
Chromatin Interaction ◽  
Similar Degree ◽  
Cell Replication ◽  
Stochastic Variation ◽  
Cell Cycles ◽  
And Function ◽  
Cell Variation

AbstractIn mammalian cells, distinct replication domains (RDs), corresponding to structural units of chromosomes called topologically-associating domains (TADs), replicate at different times during S-phase1–4. Further, early/late replication of RDs corresponds to active/inactive chromatin interaction compartments5,6. Although replication origins are selected stochastically, such that each cell is using a different cohort of origins to replicate their genomes7–12, replication-timing is regulated independently and upstream of origin selection13 and evidence suggests that replication timing is conserved in consecutive cell cycles14. Hence, quantifying the extent of cell-to-cell variation in replication timing is central to studies of chromosome structure and function. Here we devise a strategy to measure variation in single-cell replication timing using DNA copy number. We find that borders between replicated and un-replicated DNA are highly conserved between cells, demarcating active and inactive compartments of the nucleus. Nonetheless, measurable variation was evident. Surprisingly, we detected a similar degree of variation in replication timing from cell-to-cell, between homologues within cells, and between all domains genome-wide regardless of their replication timing. These results demonstrate that stochastic variation in replication timing is independent of elements that dictate timing or extrinsic environmental variation.

scholarly journals Impact of Novel Resistance Profiles in HIV-1 Reverse Transcriptase on Phenotypic Resistance to NVP

2012 ◽  
Vol 2012 ◽  
pp. 1-8
Author(s):  
Liyan Jiao ◽  
Hanping Li ◽  
Lin Li ◽  
Daomin Zhuang ◽  
Yongjian Liu ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Site Directed Mutagenesis ◽  
Drug Resistant ◽  
Wild Type ◽  
Phenotypic Resistance ◽  
Drug Resistant Mutation ◽  
The Impact ◽  
Hiv 1 ◽  
Resistant Mutation ◽  
Phenotypic Drug Resistance

Objective. To clarify the impact of H221Y mutation on drug resistance to NVP.Methods. 646 bp HIV-1polgene fragments (from 592 to 1237 nucleotide) with different NNRTIs mutation profiles from AIDS patients receiving antiretroviral therapy containing NVP regimens were introduced into pNL4-3 backbone plasmid. H221Y and (or) Y181C mutations were reverted to wild type amino acids by site-directed mutagenesis, then strains containing various mutation patterns were packaged. Phenotypic drug resistance was analyzed on TZM-bl cells.Results. 12 strains containing different drug-resistant mutation profiles were constructed, including the K101Q series (K101Q/Y181C/H221Y, K101Q/Y181C, K101Q/H221Y, and K101Q), the V179D series (V179D/Y181C/H221Y, V179D/Y181C, V179D/H221Y, and V179D), and the K103N series (K103N/Y181C/H221Y, K103N/Y181C, K103N/H221Y, K103N). For strains containing the mutation profiles (K101Q/Y181C, K101Q, V179D/Y181C, V179D, K103N/Y181C, and K103N), the presence of H221Y reduced NVP susceptibility by2.1±0.5to3.6±0.5fold. To the mutation profiles K101Q/H221Y, K101Q, V179D/H221Y, V179D, K103N/H221Y, and K103N, the presence of Y181C reduced NVP susceptibility by41.9±8.4to1297.0±289.1fold. For the strains containing K101Q, V179D, and K103N, the presence of Y181C/H221Y combination decreased NVP susceptibility by100.6±32.5to3444.6±834.5fold.Conclusion. On the bases of various NNRTIs mutation profiles, Y181C remarkably improved the IC50to NVP, although H221Ymutation alone just increases 2.1 ∼ 3.6-fold resistance to NVP, the mutation could improve 100.6 ∼ 3444.6-fold resistance to NVP when it copresent with Y181C, the phenotypic drug resistance fold was improved extremely. For strains containing the mutation profiles (K101Q/Y181C, K101Q, V179D/Y181C, V179D, K103N/Y181C, and K103N), the presence of H221Y reduced NVP susceptibility by2.1±0.5to3.6±0.5fold.

scholarly journals GWAS and functional studies implicate a role for altered DNA repair in the evolution of drug resistance in Mycobacterium tuberculosis

2022 ◽  
Author(s):  
Vinay Kumar Nandicoori ◽  
Saba Naz ◽  
Kumar Paritosh ◽  
Priyadarshini Sanyal ◽  
Sidra Khan ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Dna Repair ◽  
Mycobacterium Tuberculosis ◽  
Ex Vivo ◽  
Clinical Samples ◽  
Infection Model ◽  
Dna Repair Genes ◽  
Wild Type ◽  
The Impact ◽  
Repair Genes

The emergence of drug resistance in Mycobacterium tuberculosis (Mtb) is alarming and demands in-depth knowledge for timely diagnosis. We performed genome-wide association analysis (GWAS) using 2237 clinical strains of Mtb to identify novel genetic factors that evoke drug resistance. In addition to the known direct targets, for the first time, we identified a strong association between the mutations in the DNA repair genes and the multidrug-resistant phenotype. To evaluate the impact of variants identified in the clinical samples in the evolution of drug resistance, we utilized knockouts and complemented strains in Mycobacterium smegmatis (Msm) and Mtb. Results show that variant mutations abrogated the function of MutY and UvrB. MutY variant showed enhanced survival compared with wild-type (Rv) when the Mtb strains were subjected to multiple rounds of ex vivo antibiotic stress. Notably, in an in vivo Guinea pig infection model, the MutY variant outcompeted the wild-type strain. Collectively, we show that novel variant mutations in the DNA repair genes abrogate their function and contribute to better survival under antibiotic/host stress conditions.

scholarly journals Fitness Cost Due to Mutations in the 16S rRNA Associated with Spectinomycin Resistance in Chlamydia psittaci 6BC

2005 ◽  
Vol 49 (11) ◽  
pp. 4455-4464 ◽  
Author(s):  
Rachel Binet ◽  
Anthony T. Maurelli
Keyword(s):  
Drug Resistance ◽  
Antibiotic Resistance ◽  
Life Cycle ◽  
16S Rrna ◽  
Plaque Assay ◽  
Chlamydia Psittaci ◽  
Fitness Cost ◽  
Wild Type ◽  
Wide Range ◽  
The Impact

ABSTRACT The fitness cost of a resistance determinant is the primary parameter that determines its frequency in vivo. As a model for analysis of the impact of drug resistance mutations on the intracellular life cycle of Chlamydia spp., we studied the growth of four genetically defined spectinomycin-resistant (Spcr) clonal variants of Chlamydia psittaci 6BC isolated in the plaque assay. The development of each variant was monitored over 46 h postinfection in the absence of drug, either in pure culture or in 1:1 competition with the parent strain. Spcr mutations in the 16S rRNA gene at positions 1191 and 1193 were associated with a marked impairment of C.psittaci biological fitness, and the bacteria were severely outcompeted by the wild-type parent. In contrast, mutations at position 1192 had minor effects on the bacterial life cycle, allowing the resistant isolates to compete more efficiently with the wild-type strain. Thus, mutations with a wide range of fitness costs can be selected in the plaque assay, providing a new strategy for prediction and monitoring of the emergence of antibiotic resistance in chlamydiae. So far, drug resistance has not been a serious threat for the treatment of chlamydial infections. Tetracycline is an effective antichlamydial drug that targets 16S rRNA. Attempts to isolate spontaneous tetracycline-resistant mutants of C. psittaci 6BC revealed a frequency <3 × 10−9. We suggest that the rarity of genotypic antibiotic resistance among chlamydial clinical isolates reflects the deleterious effects of such mutations on the fitness of these obligate intracellular bacteria in the host.

Subtle Impact of Akt1 and Akt3 on Exploratory Behavior in Gene Targeted Mice

2015 ◽  
Vol 223 (3) ◽  
pp. 173-180 ◽  
Author(s):  
Christina Leibrock ◽  
Michael Hierlmeier ◽  
Undine E. Lang ◽  
Florian Lang
Keyword(s):  
Forced Swimming Test ◽  
Open Field Test ◽  
Wild Type ◽  
Average Speed ◽  
Closed Area ◽  
Light Area ◽  
Swimming Test ◽  
The Impact ◽  
Center Area ◽  
Dark Transition

Abstract. The present study explored the impact of Akt1 and Akt3 on behavior. Akt1 (akt1-/-) and Akt3 (akt3-/-) knockout mice were compared to wild type (wt) mice. The akt1-/- mice, akt3-/- mice, and wt mice were similar in most parameters of the open-field test. However, the distance traveled in the center area was slightly but significantly less in akt3-/- mice than in wt mice. In the light/dark transition test akt1-/- mice had significantly lower values than wt mice and akt3-/- mice for distance traveled, number of rearings, rearing time in the light area, as well as time spent and distance traveled in the entrance area. They were significantly different from akt3-/- mice in the distance traveled, visits, number of rearings, rearing time in the light area, as well as time spent, distance traveled, number of rearings, and rearing time in the entrance area. In the O-maze the time spent, and the visits to open arms, as well as the number of protected and unprotected headdips were significantly less in akt1-/- mice than in wt mice, whereas the time spent in closed arms was significantly more in akt1-/- mice than in wt mice. Protected and unprotected headdips were significantly less in akt3-/- mice than in wt mice. In closed area, akt3-/- mice traveled a significantly larger distance at larger average speed than akt1-/- mice. No differences were observed between akt1-/- mice, akt3-/- mice and wt-type mice in the time of floating during the forced swimming test. In conclusion, akt1-/- mice and less so akt3-/ mice display subtle changes in behavior.

scholarly journals Titanium Dioxide Presents a Different Profile in Dextran Sodium Sulphate-Induced Experimental Colitis in Mice Lacking the IBD Risk Gene Ptpn2 in Myeloid Cells

2021 ◽  
Vol 22 (2) ◽  
pp. 772
Author(s):  
Javier Conde ◽  
Marlene Schwarzfischer ◽  
Egle Katkeviciute ◽  
Janine Häfliger ◽  
Anna Niechcial ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Titanium Dioxide ◽  
Genetic Risk ◽  
Intestinal Inflammation ◽  
Sodium Sulphate ◽  
Food Additive ◽  
Wild Type ◽  
Dextran Sodium Sulphate ◽  
Risk Gene ◽  
The Impact

Environmental and genetic factors have been demonstrated to contribute to the development of inflammatory bowel disease (IBD). Recent studies suggested that the food additive; titanium dioxide (TiO2) might play a causative role in the disease. Therefore, in the present study we aimed to explore the interaction between the food additive TiO2 and the well-characterized IBD risk gene protein tyrosine phosphatase non-receptor type 2 (Ptpn2) and their role in the development of intestinal inflammation. Dextran sodium sulphate (DSS)-induced acute colitis was performed in mice lacking the expression of Ptpn2 in myeloid cells (Ptpn2LysMCre) or their wild type littermates (Ptpn2fl/fl) and exposed to the microparticle TiO2. The impact of Ptpn2 on TiO2 signalling pathways and TiO2-induced IL-1β and IL-10 levels were studied using bone marrow-derived macrophages (BMDMs). Ptpn2LysMCre exposed to TiO2 exhibited more severe intestinal inflammation than their wild type counterparts. This effect was likely due to the impact of TiO2 on the differentiation of intestinal macrophages, suppressing the number of anti-inflammatory macrophages in Ptpn2 deficient mice. Moreover, we also found that TiO2 was able to induce the secretion of IL-1β via mitogen-activated proteins kinases (MAPKs) and to repress the expression of IL-10 in bone marrow-derived macrophages via MAPK-independent pathways. This is the first evidence of the cooperation between the genetic risk factor Ptpn2 and the environmental factor TiO2 in the regulation of intestinal inflammation. The results presented here suggest that the ingestion of certain industrial compounds should be taken into account, especially in individuals with increased genetic risk

scholarly journals Baicalin Represses Type Three Secretion System of Pseudomonas aeruginosa through PQS System

Molecules ◽  
2021 ◽  
Vol 26 (6) ◽  
pp. 1497
Author(s):  
Pansong Zhang ◽  
Qiao Guo ◽  
Zhihua Wei ◽  
Qin Yang ◽  
Zisheng Guo ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Virulence Factors ◽  
Mammalian Cells ◽  
Secretion System ◽  
Chinese Herbs ◽  
Infection Model ◽  
Lung Pathology ◽  
Promising Alternative ◽  
The Impact

Therapeutics that target the virulence of pathogens rather than their viability offer a promising alternative for treating infectious diseases and circumventing antibiotic resistance. In this study, we searched for anti-virulence compounds against Pseudomonas aeruginosa from Chinese herbs and investigated baicalin from Scutellariae radix as such an active anti-virulence compound. The effect of baicalin on a range of important virulence factors in P. aeruginosa was assessed using luxCDABE-based reporters and by phenotypical assays. The molecular mechanism of the virulence inhibition by baicalin was investigated using genetic approaches. The impact of baicalin on P. aeruginosa pathogenicity was evaluated by both in vitro assays and in vivo animal models. The results show that baicalin diminished a plenty of important virulence factors in P. aeruginosa, including the Type III secretion system (T3SS). Baicalin treatment reduced the cellular toxicity of P. aeruginosa on the mammalian cells and attenuated in vivo pathogenicity in a Drosophila melanogaster infection model. In a rat pulmonary infection model, baicalin significantly reduced the severity of lung pathology and accelerated lung bacterial clearance. The PqsR of the Pseudomonas quinolone signal (PQS) system was found to be required for baicalin’s impact on T3SS. These findings indicate that baicalin is a promising therapeutic candidate for treating P. aeruginosa infections.

The impact of fusion genes on cancer stem cells and drug resistance

2021 ◽  
Author(s):  
Saurav Panicker ◽  
Sivaramakrishnan Venkatabalasubramanian ◽  
Surajit Pathak ◽  
Satish Ramalingam
Keyword(s):  
Stem Cells ◽  
Drug Resistance ◽  
Cancer Stem Cells ◽  
Fusion Genes ◽  
The Impact

scholarly journals CAND2/PMTR1 Is Required for Melatonin-Conferred Osmotic Stress Tolerance in Arabidopsis

2021 ◽  
Vol 22 (8) ◽  
pp. 4014
Author(s):  
Lin-Feng Wang ◽  
Ting-Ting Li ◽  
Yu Zhang ◽  
Jia-Xing Guo ◽  
Kai-Kai Lu ◽  
...  
Keyword(s):  
Osmotic Stress ◽  
Stress Tolerance ◽  
Wild Type Plant ◽  
Wild Type ◽  
Melatonin Receptor ◽  
Ros Scavenging ◽  
Exogenous Melatonin ◽  
Osmotic Stress Tolerance ◽  
Osmotic Stress Response ◽  
In The Wild

Osmotic stress severely inhibits plant growth and development, causing huge loss of crop quality and quantity worldwide. Melatonin is an important signaling molecule that generally confers plant increased tolerance to various environmental stresses, however, whether and how melatonin participates in plant osmotic stress response remain elusive. Here, we report that melatonin enhances plant osmotic stress tolerance through increasing ROS-scavenging ability, and melatonin receptor CAND2 plays a key role in melatonin-mediated plant response to osmotic stress. Upon osmotic stress treatment, the expression of melatonin biosynthetic genes including SNAT1, COMT1, and ASMT1 and the accumulation of melatonin are increased in the wild-type plants. The snat1 mutant is defective in osmotic stress-induced melatonin accumulation and thus sensitive to osmotic stress, while exogenous melatonin enhances the tolerance of the wild-type plant and rescues the sensitivity of the snat1 mutant to osmotic stress by upregulating the expression and activity of catalase and superoxide dismutase to repress H2O2 accumulation. Further study showed that the melatonin receptor mutant cand2 exhibits reduced osmotic stress tolerance with increased ROS accumulation, but exogenous melatonin cannot revert its osmotic stress phenotype. Together, our study reveals that CADN2 functions necessarily in melatonin-conferred osmotic stress tolerance by activating ROS-scavenging ability in Arabidopsis.

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