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acI Actinobacteria Assemble a Functional Actinorhodopsin with Natively Synthesized Retinal.

Abstract：

:Freshwater lakes harbor complex microbial communities, but these ecosystems are often dominated by acI Actinobacteria Members of this cosmopolitan lineage are proposed to bolster heterotrophic growth using phototrophy because their genomes encode actino-opsins (actR). This model has been difficult to validate experimentally because acI Actinobacteria are not consistently culturable. Based primarily on genomes from single cells and metagenomes, we provide a detailed biosynthetic route for members of acI clades A and B to synthesize retinal and its carotenoid precursors. Consequently, acI cells should be able to natively assemble light-driven actinorhodopsins (holo-ActR) to pump protons, unlike many bacteria that encode opsins but may need to exogenously obtain retinal because they lack retinal machinery. Moreover, we show that all acI clades contain genes for a secondary branch of the carotenoid pathway, implying synthesis of a complex carotenoid. Transcription analysis of acI Actinobacteria in a eutrophic lake shows that all retinal and carotenoid pathway operons are transcribed and that actR is among the most highly transcribed of all acI genes. Furthermore, heterologous expression of acI retinal pathway genes showed that lycopene, retinal, and ActR can be made using the genes encoded in these organisms. Model cells producing ActR and the key acI retinal-producing β-carotene oxygenase formed holo-ActR and acidified solution during illumination. Taken together, our results prove that acI Actinobacteria containing both ActR and acI retinal production machinery have the capacity to natively synthesize a green light-dependent outward proton-pumping rhodopsin.IMPORTANCE Microbes play critical roles in determining the quality of freshwater ecosystems, which are vital to human civilization. Because acI Actinobacteria are ubiquitous and abundant in freshwater lakes, clarifying their ecophysiology is a major step in determining the contributions that they make to nitrogen and carbon cycling. Without accurate knowledge of these cycles, freshwater systems cannot be incorporated into climate change models, ecosystem imbalances cannot be predicted, and policy for service disruption cannot be planned. Our work fills major gaps in microbial light utilization, secondary metabolite production, and energy cycling in freshwater habitats.

journal_name

Appl Environ Microbiol

journal_title

Applied and environmental microbiology

authors

Dwulit-Smith JR,Hamilton JJ,Stevenson DM,He S,Oyserman BO,Moya-Flores F,Garcia SL,Amador-Noguez D,McMahon KD,Forest KT

doi

10.1128/AEM.01678-18

subject

Has Abstract

pub_date

2018-11-30 00:00:00

issue

eissn

0099-2240

issn

1098-5336

pii

AEM.01678-18

journal_volume

pub_type

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acI Actinobacteria Assemble a Functional Actinorhodopsin with Natively Synthesized Retinal.