Epistatic and Synergistic Interactions Between Circadian Clock Mutations in Neurospora crassa

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.

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Isolation and Characterization of a Temperature-Sensitive Circadian Clock Mutant of Neurospora crassa

Genetics ◽

10.1093/genetics/146.2.525 ◽

1997 ◽

Vol 146 (2) ◽

pp. 525-530 ◽

Cited By ~ 2

Author(s):

Louis W Morgan ◽

Jerry F Feldman

Keyword(s):

Circadian Clock ◽

Neurospora Crassa ◽

Mutant Strain ◽

Double Mutant ◽

Period Length ◽

Temperature Sensitive ◽

Wild Type ◽

Isolation And Characterization ◽

Double Mutant Strain ◽

Clock Mutant

A new circadian clock mutant has been isolated in Neurospora crassa. This new mutation, called period-6 (pd-6), has two features novel to known clock mutations. First, the mutation is temperature sensitive. At restrictive temperatures (above 21°) the mutation shortens circadian period length from a wild-type value of 21.5 hr to 18 hr. At permissive temperatures (below 21°) the mutant has a 20.5-hr period length close to that of the wild-type strain. Second, the prd-6 mutation is epistatic to the previously isolated clock mutation period-2 (prd-2). This epistasis is unusual in that the prd-2 prd-6 double mutant strain has an 18-hr period length at both the restrictive and permissive temperatures. That is, the temperature-sensitive aspect of the phenotype of the prd-6 strain is lost in the prd-2 prd-6 double mutant strain. This suggests that the gene products of the prd-2 and prd-6 loci may interact physically and that the presence of a normal prd-2+ protein is required for low temperature to “rescue” the prd-6 mutant phenotype. These results, combined with our recent finding that prd-2 and some alleles of the frq gene show genetic synergy, suggest that it may be possible to establish a more comprehensive model of the Neurospora circadian clock.

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GENETIC AND PHYSIOLOGICAL CHARACTERISTICS OF A SLOW-GROWING CIRCADIAN CLOCK MUTANT OF NEUROSPORA CRASSA

Genetics ◽

10.1093/genetics/88.2.255 ◽

1978 ◽

Vol 88 (2) ◽

pp. 255-265

Author(s):

Jerry F Feldman ◽

Cheryl A Atkinson

Keyword(s):

Linkage Group ◽

Circadian Clock ◽

Neurospora Crassa ◽

Double Mutant ◽

Slow Growth ◽

Period Length ◽

The Other ◽

Wild Type ◽

Long Period ◽

Clock Mutant

ABSTRACT A circadian clock mutant of Neurospora crassa with a period length of about 25.8 hours (4 hr longer than wild type) has been isolated after mutagenesis of the band strain. This mutant, called frq-5, segregates as a single nuclear gene, maps near the centromere on linkage group III, and is unlinked to four previously described clock mutants clustered on linkage group VII R (Feldman and Hoyle 1973, 1976). frq-5 differs from the other clock mutants in at least two other respects: (1) it is recessive in heterokaryons, and (2) it grows at about 60% the rate of the parent band strain on both minimal and complete media. Double mutants between frq-5 and each of the other clock mutants show additivity of period length-two long period mutants produce a double mutant whose period length is longer than either of the two single mutants, while a long and a short period double mutant has an intermediate period length. Although slow growth and long periodicity of frq-5 have segregated together among more than 300 progeny, slow growth per se is not responsible for the long period, since all the double mutants have the slow growth characteristic of frq-5, but have period lengths both shorter and longer than wild type.

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COMPLEMENTATION ANALYSIS OF METABOLITE-RESISTANT MUTATIONS WITH FORCED HETEROKARYONS OF NEUROSPORA CRASSA

Genetics ◽

10.1093/genetics/74.4.581 ◽

1973 ◽

Vol 74 (4) ◽

pp. 581-593

Author(s):

W M Thwaites ◽

F K Knauert ◽

S S Carney

Keyword(s):

Positive Result ◽

Neurospora Crassa ◽

Mutant Strain ◽

Uptake Rate ◽

Double Mutant ◽

Complementation Test ◽

Gene Products ◽

Double Mutant Strain ◽

Common Precursor ◽

Resistant Mutants

ABSTRACT The double mutant strain pyr-3 arg-12s is a prototroph because a common precursor of arginine and pyrimidine is supplied by the arginine pathway. Growth of this strain is inhibited by exogenous citrulline or arginine. Citrulline-resistant mutants of this strain were selected, and they resulted from modifier mutations at other loci. Forced heterokaryons were used to study complementation among these modifiers. Since the complementation test requires the scoring of non-growth as the positive result, there was concern that variations in nuclear ratios could give erroneous results. This possibility does not seem significant, since groups of mutants established by complementation correspond with groups established by physiological, enzymatic, and recombinational measurements.—The technique has revealed that the most frequently mutated loci are arg-1 and what is probably un-3. Arg-1 mutations affect the conversion of citrulline to argininosuccinate, while un-3 mutations reduce the citrulline uptake rate. Since most of these mutations are of the intracistronic complementing type, a complementation map was constructed for most of the affected loci. The high proportion of complementors in each map can be explained by assuming that partially functioning gene products are more likely to complement with each other than are those which are nonfunctional.

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

Advanced Research on Plant Lipids ◽

10.1007/978-94-017-0159-4_14 ◽

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

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

Frontiers in Microbiology ◽

10.3389/fmicb.2021.659233 ◽

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.

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

Eukaryotic Cell ◽

10.1128/ec.00237-10 ◽

2011 ◽

Vol 10 (7) ◽

pp. 869-883 ◽

Cited By ~ 4

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.

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

Organic Chemistry Frontiers ◽

10.1039/c5qo00433k ◽

2016 ◽

Vol 3 (4) ◽

pp. 496-500 ◽

Cited By ~ 11

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.

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Distinct Signaling Pathways from the Circadian Clock Participate in Regulation of Rhythmic Conidiospore Development in Neurospora crassa

Eukaryotic Cell ◽

10.1128/ec.1.2.273-280.2002 ◽

2002 ◽

Vol 1 (2) ◽

pp. 273-280 ◽

Cited By ~ 20

Author(s):

Alejandro Correa ◽

Deborah Bell-Pedersen

Keyword(s):

Gene Expression ◽

Circadian Clock ◽

Neurospora Crassa ◽

Mutant Strain ◽

Specific Gene ◽

Developmentally Regulated ◽

Environmental Signals ◽

Endogenous Clock ◽

Environmental Induction ◽

High Level

ABSTRACT Several different environmental signals can induce asexual spore development (conidiation) and expression of developmentally regulated genes in Neurospora crassa. However, under constant conditions, where no environmental cues for conidiation are present, the endogenous circadian clock in N. crassa promotes daily rhythms in expression of known developmental genes and of conidiation. We anticipated that the same pathway of gene regulation would be followed during clock-controlled conidiation and environmental induction of conidiation and that the circadian clock would need only to control the initial developmental switch. Previous experiments showed that high-level developmental induction of the clock-controlled genes eas (ccg-2) and ccg-1 requires the developmental regulatory proteins FL and ACON-2, respectively, and normal developmental induction of fl mRNA expression requires ACON-2. We demonstrate that the circadian clock regulates rhythmic fl gene expression and that fl rhythmicity requires ACON-2. However, we find that clock regulation of eas (ccg-2) is normal in an fl mutant strain and ccg-1 expression is rhythmic in an acon-2 mutant strain. Together, these data point to the endogenous clock and the environment following separate pathways to regulate conidiation-specific gene expression.

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

Journal of Bacteriology ◽

10.1128/jb.01005-12 ◽

2012 ◽

Vol 194 (23) ◽

pp. 6398-6409 ◽

Cited By ~ 45

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.

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