Specific mutations in the permease domain of septal protein SepJ differentially affect functions related to multicellularity in the filamentous cyanobacterium Anabaena.

Félix Ramos-León,Vicente Mariscal,Enrique Flores,Sergio Arévalo

doi:10.15698/mic2018.12.661

Abstract

Filamentous, heterocyst-forming cyanobacteria are multicellular organisms in which growth requires the activity of two interdependent cell types that exchange nutrients and regulators. Vegetative cells provide heterocysts with reduced carbon, and heterocysts provide vegetative cells with fixed nitrogen. Additionally, heterocyst differentiation from vegetative cells is regulated by inhibitors of differentiation produced by prospective heterocysts and heterocysts. Proteinaceous structures known as septal junctions join the cells in the filament. The SepJ protein is involved in formation of septal junctions in the model heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120. SepJ bears extra-membrane and membrane (permease) domains and is located at the cell poles in the intercellular septa of the filament. Here we created Anabaena mutants that produce SepJ proteins altered in the permease domain. Some of these mutant SepJ proteins did not provide functions needed for Anabaena to form long filaments and (in some cases) differentiate heterocysts, identifying amino acids and amino acid stretches that are important for the structure or function of the protein. Some other mutant SepJ proteins fulfilled filamentation and heterocyst differentiation functions but failed to provide normal communication function assessed via the intercellular transfer of the fluorescent marker calcein. These mutant SepJ proteins bore mutations in amino acids located at the cytoplasmic face of the permease, which could affect access of the fluorescent marker to the septal junctions. Overall, the data are consistent with the idea that SepJ carries out multiple roles in the multicellular function of the Anabaena filament.

Highlights

Heterocyst-forming cyanobacteria grow as chains of vegetative cells that fix CO2, performing oxygenic photosynthesis
As an important contributor to the formation of septal junctions in Anabaena, the SepJ protein is needed to make long filaments in both media containing and media lacking a source of combined nitrogen, its effect is most stringent in the absence of combined nitrogen [18, 19]
SepJ is required for Anabaena to make a normal number of septal peptidoglycan nanopores and is involved in intercellular molecular exchange as shown with fluorescent markers, especially calcein [7, 9, 24]

Summary

Introduction

Heterocyst-forming cyanobacteria grow as chains of vegetative cells that fix CO2, performing oxygenic photosynthesis. Strain PCC 7120 (hereafter Anabaena), differentiation requires the activity of the NtcA and HetR transcription factors [2], and differentiation of an excessive number of heterocysts appears to be prevented by diffusible inhibitors produced by prospective heterocysts (proheterocysts) and heterocysts [1]. This inhibition involves possible morphogens related to products of the patS and hetN genes [3,4,5]. Exchanged nutrients likely include sucrose transferred from vegetative cells to heterocysts (see [7], and references therein), and glutamine and the dipeptide β-aspartyl arginine transferred from heterocysts to vegetative cells (see [8], and references therein)

Methods

Results

Conclusion