scholarly journals Role of Hsl7 in Morphology and Pathogenicity and Its Interaction with Other Signaling Components in the Plant Pathogen Ustilago maydis

2011 ◽  
Vol 10 (7) ◽  
pp. 869-883 ◽  
Author(s):  
C. Ben Lovely ◽  
Kavita Burman Aulakh ◽  
Michael H. Perlin
Keyword(s):  
Ammonium Sulfate ◽  
Mutant Strain ◽  
Double Mutant ◽  
Ustilago Maydis ◽  
Cell Length ◽  
Phytopathogenic Fungus ◽  
Nitrogen Base ◽  
Content Type ◽  
Dimorphic Transition ◽  
Double Mutant Strain

ABSTRACTThe phytopathogenic fungusUstilago maydisundergoes a dimorphic transition in response to mating pheromone, host, and environmental cues. On a solid medium deficient in ammonium (SLAD [0.17% yeast nitrogen base without ammonium sulfate or amino acids, 2% dextrose, 50 μM ammonium sulfate]),U. maydisproduces a filamentous colony morphology, while in liquid SLAD, the cells do not form filaments. The p21-activated protein kinases (PAKs) play a substantial role in regulating the dimorphic transition in fungi. The PAK-like Ste20 homologue Smu1 is required for a normal response to pheromone, via upregulation of pheromone expression, and virulence, and its disruption affects both processes. Our experiments suggest that Smu1 also regulates cell length and the filamentous response on solid SLAD medium. Yeast two-hybrid analysis suggested an Hsl7 homologue as a potential interacting partner of Smu1, and a unique open reading frame for such an arginine methyltransferase was detected in theU. maydisgenome sequence. Hsl7 regulates cell length and the filamentous response to solid SLAD in a fashion opposite to that of Smu1, but neither overexpression nor disruption ofhsl7attenuates virulence. Simultaneous disruption ofhsl7and overexpression ofsmu1lead to a hyperfilamentous response on solid SLAD. Moreover, only this double mutant strain forms filaments in liquid SLAD. The double mutant strain was also significantly reduced in virulence. A similar filamentous response in both solid and liquid SLAD was observed in strains lacking another PAK-like protein kinase involved in cytokinesis and polar growth, Cla4. Our data suggest that Hsl7 may regulate cell cycle progression, while both Smu1 and Cla4 appear to be involved in the filamentous response inU. maydis.

scholarly journals Mycobacterium tuberculosis CwsA Interacts with CrgA and Wag31, and the CrgA-CwsA Complex Is Involved in Peptidoglycan Synthesis and Cell Shape Determination

2012 ◽  
Vol 194 (23) ◽  
pp. 6398-6409 ◽  
Author(s):  
P. Plocinski ◽  
N. Arora ◽  
K. Sarva ◽  
E. Blaszczyk ◽  
H. Qin ◽  
...  
Keyword(s):  
Cell Wall ◽  
Mycobacterium Tuberculosis ◽  
Mutant Strain ◽  
Cell Shape ◽  
Double Mutant ◽  
Bacterial Cell Division ◽  
Cell Wall Synthesis ◽  
Content Type ◽  
Ring Assembly ◽  
Double Mutant Strain

ABSTRACTBacterial cell division and cell wall synthesis are highly coordinated processes involving multiple proteins. Here, we show that Rv0008c, a novel small membrane protein fromMycobacterium tuberculosis, localizes to the poles and on membranes and shows an overall punctate localization throughout the cell. Furthermore, Rv0008c interacts with two proteins, CrgA and Wag31, implicated in peptidoglycan (PG) synthesis in mycobacteria. Deletion of the Rv0008c homolog inM. smegmatis, MSMEG_0023, caused bulged cell poles, formation of rounded cells, and defects in polar localization of Wag31 and cell wall synthesis, with cell wall synthesis measured by the incorporation of the [14C]N-acetylglucosamine cell wall precursor. TheM. smegmatisMSMEG_0023crgAdouble mutant strain showed severe defects in growth, viability, cell wall synthesis, cell shape, and the localization of the FtsZ, FtsI, and Wag31 proteins. The double mutant strain also exhibited increased autolytic activity in the presence of detergents. Because CrgA and Wag31 proteins interact with FtsI individually, we believe that regulated cell wall synthesis and cell shape maintenance require the concerted actions of the CrgA, Rv0008c, FtsI, and Wag31 proteins. We propose that, together, CrgA and Rv0008c, renamed CwsA forcellwall synthesis and cellshape proteinA, play crucial roles in septal and polar PG synthesis and help coordinate these processes with the FtsZ-ring assembly in mycobacteria.

scholarly journals Epistatic and Synergistic Interactions Between Circadian Clock Mutations in Neurospora crassa

Genetics ◽  
2001 ◽  
Vol 159 (2) ◽  
pp. 537-543
Author(s):  
Louis W Morgan ◽  
Jerry F Feldman
Keyword(s):  
Circadian Clock ◽  
Neurospora Crassa ◽  
Mutant Strain ◽  
Temperature Compensation ◽  
Double Mutant ◽  
Synergistic Interactions ◽  
Long Period ◽  
Double Mutant Strain ◽  
Short Period ◽  
Compensation Mechanisms

Abstract We identified a series of epistatic and synergistic interactions among the circadian clock mutations of Neurospora crassa that indicate possible physical interactions among the various clock components encoded by these genes. The period-6 (prd-6) mutation, a short-period temperature-sensitive clock mutation, is epistatic to both the prd-2 and prd-3 mutations. The prd-2 and prd-3 long-period mutations show a synergistic interaction in that the period length of the double mutant strain is considerably longer than predicted. In addition, the prd-2 prd-3 double mutant strain also exhibits overcompensation to changes in ambient temperature, suggesting a role in the temperature compensation machinery of the clock. The prd-2, prd-3, and prd-6 mutations also show significant interactions with the frq7 long-period mutation. These results suggest that the gene products of prd-2, prd-3, and prd-6 play an important role in both the timing and temperature compensation mechanisms of the circadian clock and may interact with the FRQ protein.

Characterizing KAS Enzymes Using the E. Coli Double Mutant Strain CY244

2003 ◽  
pp. 65-68
Author(s):  
Anne Vinther Rasmussen ◽  
Rie Yasuno ◽  
Penny von Wettstein-Knowles
Keyword(s):  
Mutant Strain ◽  
Double Mutant ◽  
E Coli ◽  
Double Mutant Strain

scholarly journals Lack of the Bacterial Phytochrome Protein Decreases Deinococcus radiodurans Resistance to Mitomycin C

2021 ◽  
Vol 12 ◽  
Author(s):  
Jong-Hyun Jung ◽  
Soyoung Jeong ◽  
Seonghun Im ◽  
Min-Kyu Kim ◽  
Ho Seong Seo ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Survival ◽  
Mitomycin C ◽  
Mutant Strain ◽  
Double Mutant ◽  
Deinococcus Radiodurans ◽  
Red Light ◽  
Double Mutant Strain ◽  
Significant Difference

Deinococcus radiodurans known for its extraordinary resistance to ionizing radiation contains bacterial phytochrome (BphP), a member of the family of red/far-red light-sensing proteins. In this study, we constructed a bphP mutant strain (ΔbphP) to investigate the role of D. radiodurans BphP (DrBphP) in the DNA damage response. When cells were incubated under light and dark conditions following exposure to DNA damaging agents, such as γ- and UV-radiation and mitomycin C (MMC), no significant difference in cell survival was observed between the wild-type D. radiodurans strain (WT) and ΔbphP. However, when continuously exposed to MMC under light conditions, the WT strain notably exhibited increased survival compared to cells grown in the dark. The increased survival was not observed in the ΔbphP strain. These results are indicative of the protective role of light-activated DrBphP in the presence of MMC. Site-directed mutagenesis revealed that the conserved amino acids Cys-24 and His-532 involved in chromophore binding and signal transduction, respectively, were essential for the protective function of DrBphP. Inactivation of the cognate response regulator (RR; DrBphR) of DrBphP increased MMC resistance in the dark. In trans complementation of the bphP bphR double mutant strain (ΔbphPR) with DrBphR decreased MMC resistance. Considering that DrBphP acts as a light-activated phosphatase that dephosphorylates DrBphR, it appears that phosphorylated DrBphR exerts a negative effect on cell survival in the presence of MMC. DrBphP overexpression resulted in an increase in MMC resistance of ΔbphPR, implying that other RRs might be involved in the DrBphP-mediated signaling pathway. A mutant lacking the dr_0781 gene (Δdr_0781) demonstrated the same MMC phenotype as ΔbphR. Survival was further increased in the bphR dr_0781 double mutant strain compared to each single mutant ΔbphR or Δdr_0781, suggesting that DR_0781 is also involved in the DrBphP-dependent MMC sensitivity. This study uncovered a previously unknown phenomenon of red/far-red light-dependent DNA damage survival mediated by BphP by identifying the conditions under which DrBphP exhibits a fitness advantage.

Concurrent modifications of the C-terminus and side ring of thiostrepton and their synergistic effects with respect to improving antibacterial activities

2016 ◽  
Vol 3 (4) ◽  
pp. 496-500 ◽  
Author(s):  
Shoufeng Wang ◽  
Qingfei Zheng ◽  
Jianfeng Wang ◽  
Dandan Chen ◽  
Yunsong Yu ◽  
...  
Keyword(s):  
Mutant Strain ◽  
Double Mutant ◽  
Synergistic Effects ◽  
Antibacterial Activities ◽  
Ring Structure ◽  
Quinaldic Acid ◽  
C Terminus ◽  
Double Mutant Strain

Five new C-terminally methylated TSR derivatives that varied in side-ring structure were obtained via the chemical feeding of quinaldic acid analogs to a double-mutant strain ΔtsrB/T.

scholarly journals Disruption of the cpsE and endA Genes Attenuates Streptococcus pneumoniae Virulence: Towards the Development of a Live Attenuated Vaccine Candidate

Vaccines ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 187 ◽  
Author(s):  
Malik Amonov ◽  
Nordin Simbak ◽  
Wan Mohd. Razin Wan Hassan ◽  
Salwani Ismail ◽  
Nor Iza A. Rahman ◽  
...  
Keyword(s):  
Developing Countries ◽  
Streptococcus Pneumoniae ◽  
Mutant Strain ◽  
Double Mutant ◽  
Gene Knockout ◽  
High Serum ◽  
High Dose ◽  
Wild Type ◽  
Knockout Mutant ◽  
Double Mutant Strain

The majority of deaths due to Streptococcus pneumoniae infections are in developing countries. Although polysaccharide-based pneumococcal vaccines are available, newer types of vaccines are needed to increase vaccine affordability, particularly in developing countries, and to provide broader protection across all pneumococcal serotypes. To attenuate pneumococcal virulence with the aim of engineering candidate live attenuated vaccines (LAVs), we constructed knockouts in S. pneumoniae D39 of one of the capsular biosynthetic genes, cpsE that encodes glycosyltransferase, and the endonuclease gene, endA, that had been implicated in the uptake of DNA from the environment as well as bacterial escape from neutrophil-mediated killing. The cpsE gene knockout significantly lowered peak bacterial density, BALB/c mice nasopharyngeal (NP) colonisation but increased biofilm formation when compared to the wild-type D39 strain as well as the endA gene knockout mutant. All constructed mutant strains were able to induce significantly high serum and mucosal antibody response in BALB/c mice. However, the cpsE-endA double mutant strain, designated SPEC, was able to protect mice from high dose mucosal challenge of the D39 wild-type. Furthermore, SPEC showed 23-fold attenuation of virulence compared to the wild-type. Thus, the cpsE-endA double-mutant strain could be a promising candidate for further development of a LAV for S. pneumoniae.

scholarly journals An msbB Homologue Carried in Plasmid pO157 Encodes an Acyltransferase Involved in Lipid A Biosynthesis in Escherichia coli O157:H7

2004 ◽  
Vol 72 (2) ◽  
pp. 1174-1180 ◽  
Author(s):  
Sang-Hyun Kim ◽  
Wenyi Jia ◽  
Russell E. Bishop ◽  
Carlton Gyles
Keyword(s):  
Escherichia Coli ◽  
Mutant Strain ◽  
Double Mutant ◽  
Lipid A ◽  
Escherichia Coli O157 ◽  
Wild Type ◽  
Single Mutant ◽  
Double Mutant Strain ◽  
Mutant Strains ◽  
Lipid A Biosynthesis

ABSTRACT Escherichia coli O157:H7 carries a chromosomal msbB1 and a plasmid-encoded msbB2 gene. We characterized msbB2 function as a homologue of msbB1 by examination of wild-type organisms and mutant strains that lacked functional msbB1, msbB2, and both msbB1 and msbB2. The msbB double-mutant strain generated pentaacyl lipid A, while the single-mutant strains synthesized hexaacyl lipid A. Complementation with overexpressed msbB2 converted pentaacyl into hexaacyl lipid A in the double-mutant strain. The transcription of both msbB genes occurred simultaneously. Lack of MsbB2 activity slightly increased the microheterogeneity of the lipid A species. These results suggest that the msbB2 gene plays a role not only in the routine generation of fully hexaacylated lipid A but also in suppressing the microheterogeneity of lipid A species, the endotoxic determinant of the organism.

scholarly journals Cryptococcus neoformansGlucuronoxylomannan and Sterylglucoside Are Required for Host Protection in an Animal Vaccination Model

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
Ana Caroline Colombo ◽  
Antonella Rella ◽  
Tyler Normile ◽  
Luna S. Joffe ◽  
Patricia M. Tavares ◽  
...  
Keyword(s):  
Double Mutant ◽  
Capsular Polysaccharide ◽  
Galleria Mellonella ◽  
Genetic Manipulation ◽  
Vaccination Strategy ◽  
Gene Encoding ◽  
Content Type ◽  
Host Protection ◽  
Double Mutant Strain ◽  
Animal Vaccination

ABSTRACTCryptococcus neoformansis an encapsulated fungal pathogen that causes meningoencephalitis. There are no prophylactic tools for cryptococcosis. Previously, our group showed that aC. neoformansmutant lacking the gene encoding sterylglucosidase (Δsgl1) induced protection in both immunocompetent and immunocompromised murine models of cryptococcosis. Since sterylglucosidase catalyzes degradation of sterylglucosides (SGs), accumulation of this glycolipid could be responsible for protective immunity. In this study, we analyzed whether the activity of SGs is sufficient for the protective effect induced by the Δsgl1strain. We observed that the accumulation of SGs impacted several properties of the main polysaccharide that composes the fungal capsule, glucuronoxylomannan (GXM). We therefore used genetic manipulation to delete theSGL1gene in the acapsular mutant Δcap59to generate a double mutant (strain Δcap59/Δsgl1) that was shown to be nonpathogenic and cleared from the lung of mice within 7 days post-intranasal infection. The inflammatory immune response triggered by the Δcap59/Δsgl1mutant in the lung differed from the response seen with the other strains. The double mutant did not induce protection in a vaccination model, suggesting that SG-related protection requires the main capsular polysaccharide. Finally, GXM-containing extracellular vesicles (EVs) enriched in SGs delayed the acute lethality ofGalleria mellonellaagainstC. neoformansinfection. These studies highlighted a key role for GXM and SGs in inducing protection against a secondary cryptococcal infection, and, since EVs notoriously contain GXM, these results suggest the potential use of Δsgl1EVs as a vaccination strategy for cryptococcosis.IMPORTANCEThe number of deaths from cryptococcal meningitis is around 180,000 per year. The disease is the second leading cause of mortality among individuals with AIDS. Antifungal treatment is costly and associated with adverse effects and resistance, evidencing the urgency of development of both therapeutic and prophylactic tools. Here we demonstrate the key roles of polysaccharide- and glycolipid-containing structures in a vaccination model to prevent cryptococcosis.

Sign in / Sign up

Export Citation Format

Share Document