The blue light-dependent LOV-protein LdaP of Dinoroseobacter shibae acts as antirepressor of the PpsR repressor, regulating photosynthetic gene cluster expression

Saskia Pucelik; Miriam Becker; Steffi Heyber; Lars Wöhlbrand; Ralf Rabus; Dieter Jahn; Elisabeth Härtig

doi:10.3389/fmicb.2024.1351297

The blue light-dependent LOV-protein LdaP of Dinoroseobacter shibae acts as antirepressor of the PpsR repressor, regulating photosynthetic gene cluster expression

Front Microbiol. 2024 Feb 7:15:1351297. doi: 10.3389/fmicb.2024.1351297. eCollection 2024.

Authors

Saskia Pucelik¹, Miriam Becker¹, Steffi Heyber¹, Lars Wöhlbrand², Ralf Rabus², Dieter Jahn³, Elisabeth Härtig¹

Affiliations

¹ Institute of Microbiology, Technische Universität Braunschweig, Braunschweig, Germany.
² Institute for Chemistry and Biology of the Marine Environment (ICBM), Carl von Ossietzky University of Oldenburg, Oldenburg, Germany.
³ Braunschweig Integrated Centre of Systems Biology (BRICS), Technische Universität Braunschweig, Braunschweig, Germany.

Abstract

In the marine α-proteobacterium Dinoroseobacter shibae more than 40 genes of the aerobic anoxygenic photosynthesis are regulated in a light-dependent manner. A genome-wide screen of 5,605 clones from a D. shibae transposon library for loss of pigmentation and changes in bacteriochlorophyll absorbance identified 179 mutant clones. The gene encoding the LOV-domain containing protein Dshi_1135 was identified by its colorless phenotype. The mutant phenotype was complemented by the expression of a Dshi_1135-strep fusion protein in trans. The recombinantly produced and chromatographically purified Dshi_1135 protein was able to undergo a blue light-induced photocycle mediated by bound FMN. Transcriptome analyses revealed an essential role for Dshi_1135 in the light-dependent expression of the photosynthetic gene cluster. Interactomic studies identified the repressor protein PpsR as an interaction partner of Dshi_1135. The physical contact between PpsR and the Dshi_1135 protein was verified in vivo using the bacterial adenylate cyclase-based two-hybrid system. In addition, the antirepressor function of the Dshi_1135 protein was demonstrated in vivo testing of a bchF-lacZ reporter gene fusion in a heterologous Escherichia coli-based host system. We therefore propose to rename the Dshi_1135 protein to LdaP (light-dependent antirepressor of PpsR). Using the bacterial two-hybrid system, it was also shown that cobalamin (B₁₂) is essential for the interaction of the antirepressor PpaA with PpsR. A regulatory model for the photosynthetic gene cluster in D. shibae was derived, including the repressor PpsR, the light-dependent antirepressor LdaP and the B₁₂-dependent antirepressor PpaA.

Keywords: Dinoroseobacter shibae; LOV; Roseobacter; aerobic anoxygenic photosynthesis; blue light-dependent gene regulation; photosynthetic gene cluster; protein–protein interaction; transposon library screen.

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was funded by the Deutsche Forschungsgemeinschaft (DFG) in the framework of the Transregional Collaborative Researche Centre (TRR51): Ecology, Physiology and Molecular Biology of the Roseobacter Group.