A Repressor-Antirepressor Pair Links Two Loci Controlling Light-induced Carotenogenesis in Myxococcus xanthus

Lopez-Rubio, José-Juan; Elías‐Arnanz, Montserrat; Padmanabhan, S.; Murillo, Francisco J.

doi:10.1074/jbc.m110351200

Cited by 31 publications

(85 citation statements)

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“…Overexpression and Purification of His-tagged CarA and CarSPlasmid constructs for overexpressing His 6 -tagged CarA and CarS, their overexpression, purification, and concentration determination have been described previously (22). His 6 -CarS was expressed as a soluble protein and purified under native conditions.…”

Section: Methodsmentioning

confidence: 99%

“…Gel Mobility Shift and DNase I Footprinting Assays-PCRs with one of the two primers radiolabeled at the 5Ј-end were carried out to generate the DNA probes (22). Probes with specific mutations were obtained by the PCR overlap extension method (36).…”

Section: Methodsmentioning

confidence: 99%

“…The photo-induced expression of the carB operon is regulated by CarA, which is produced independently of light from an adjacent operon, and by CarS (28,29). In the dark, carB is repressed by the sequence-specific binding of CarA to a DNA stretch that includes an interrupted palindrome located upstream of the carB promoter (P B ) (22). Illumination leads to an increase in the levels of CarS, which physically interacts with CarA, and thereby dismantles the CarA⅐DNA complex to cause derepression of carB (21,22).…”

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confidence: 99%

“…In the dark, carB is repressed by the sequence-specific binding of CarA to a DNA stretch that includes an interrupted palindrome located upstream of the carB promoter (P B ) (22). Illumination leads to an increase in the levels of CarS, which physically interacts with CarA, and thereby dismantles the CarA⅐DNA complex to cause derepression of carB (21,22). Our previous study identified the palindrome that is present upstream of P B (between positions Ϫ47 and Ϫ64 relative to the transcription start point) as an essential element of the CarA operator.…”

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confidence: 99%

“…carF is a recently identified gene whose product has been proposed to function as an anti-anti-factor at the earliest step in the regulatory cascade (17). The carQRS locus is a key regulatory operon that encodes three proteins with low, basal levels of expression in the dark: the extracytoplasmic function -factor CarQ (18 -20), its antifactor CarR (18,20), and the antirepressor protein CarS (21,22). Central to the induction of carotenogenesis on exposure to light is the CarF-mediated inactivation of CarR to release CarQ (17,18).…”

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confidence: 99%

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Operator Design and Mechanism for CarA Repressor-mediated Down-regulation of the Photoinducible carB Operon in Myxococcus xanthus

Lopez-Rubio

Padmanabhan

Lázaro

et al. 2004

Journal of Biological Chemistry

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The carB operon encodes all except one of the enzymes involved in light-induced carotenogenesis in Myxococcus xanthus. Expression of its promoter (P B ) is repressed in the dark by sequence-specific DNA binding of CarA to a palindrome (pI) located between positions ؊47 and ؊64 relative to the transcription start site. This promotes subsequent binding of CarA to additional sites that remain to be defined. CarS, produced in the light, interacts physically with CarA, abrogates CarA-DNA binding, and thereby derepresses P B . In this study, we delineate the operator design that exists for CarA by precisely mapping out the second operator element. For this, we examined how stepwise deletions and site-directed mutagenesis in the region between the palindrome and the transcription start site affect CarA binding around P B in vitro and expression of P B in vivo. These revealed the second operator element to be an imperfect interrupted palindrome (pII) spanning positions ؊26 to ؊40. In vitro assays using purified M. xanthus RNA polymerase showed that CarA abolishes P B -RNA polymerase binding and runoff transcription and that both were restored by CarS, thus rationalizing the observations in vivo. CarA binding to pII (after association with pI) effectively occludes RNA polymerase from P B and so provides the operative mechanism for the repression of the carB operon by CarA. The bipartite operator design, whereby transcription is blocked by the low affinity CarA-pII binding and is readily restored by CarS, may have evolved to match the needs for a rapid and an effective response to light.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Operator Design and Mechanism for CarA Repressor-mediated Down-regulation of the Photoinducible carB Operon in Myxococcus xanthus

Lopez-Rubio

Padmanabhan

Lázaro

et al. 2004

Journal of Biological Chemistry

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Light Mediation as a Strategy to Induce Production of Valuable Microbial Compounds

Gan

Ting

2019

Microorganisms for Sustainability

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Conversion of the lycopene monocyclase of Myxococcus xanthus into a bicyclase

Iniesta

Cervantes

Murillo

2008

Appl Microbiol Biotechnol

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Depending on the cyclized hydrocarbon backbone ends, carotenoids can be acyclic, monocyclic, or bicyclic. Lycopene cyclases are the enzymes responsible for catalyzing the formation of cyclic carotenoids from acyclic lycopene. Myxococcus xanthus is a bacterium that accumulates monocyclic carotenoids such as a glycoside ester of myxobacton. We show here that this bacterium possesses a cyclase belonging to the group of the heterodimeric cyclases CrtYc and CrtYd. These two individual proteins are encoded by crtYc and crtYd, which are located in the carotenogenic carA operon of the carB-carA gene cluster, and the presence of both is essential for the cyclization of lycopene. CrtYc and CrtYd from M. xanthus form a heterodimeric cyclase with beta-monocyclic activity, which converts lycopene into monocyclic gamma-carotene, but not into bicyclic beta-carotene like most beta-cyclases. This is an unusual case where two different proteins constitute a lycopene cyclase enzyme with monocyclic activity. We were able to convert this lycopene monocyclase into a lycopene bicyclase enzyme producing beta-carotene, by fusing both proteins with an extra transmembrane domain. The chimeric protein appears to allow a proper membranal disposition of both CrtYc and CrtYd, to perform two cyclization reactions, while a hybrid without the extra transmembrane helix performs only one cyclization.

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A Repressor-Antirepressor Pair Links Two Loci Controlling Light-induced Carotenogenesis in Myxococcus xanthus

Cited by 31 publications

References 35 publications

Operator Design and Mechanism for CarA Repressor-mediated Down-regulation of the Photoinducible carB Operon in Myxococcus xanthus

Operator Design and Mechanism for CarA Repressor-mediated Down-regulation of the Photoinducible carB Operon in Myxococcus xanthus

Light Mediation as a Strategy to Induce Production of Valuable Microbial Compounds

Conversion of the lycopene monocyclase of Myxococcus xanthus into a bicyclase

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