Document Type

Dissertation - Open Access

Award Date

2018

Degree Name

Doctor of Philosophy (PhD)

Department / School

Biology and Microbiology

First Advisor

Ruanbao Zhou

Keywords

Akinetes, Cyanobacteria, Heterocyst genome, Lectins Oxic, Nitrogen fixation, Proteomics

Abstract

Cyanobacteria are gram-negative photosynthetic bacteria. In some filamentous cyanobacteria such as Anabaena cylindrica, every 10th to 20th vegetative cells can differentiate into nitrogen fixing heterocysts. Heterocyst can induce the neighboring cells developing into spore-like akinetes. The specialized cell functions and resourceful networks have contributed to the prosperity of cyanobacteria for over 2 billion years, but the genetic mechanisms for multi-cellular differentiation are barely known, especially for akinete formation. The heterocysts, akinetes and vegetative cells of Anabaena cylindrica were isolated for proteomic study. This study identified a total of 1395 proteins, including 664 proteins from akinetes, 751 proteins from heterocysts, and 1236 proteins from vegetative cells. There were 45 proteins (33 novel proteins) found exclusive in akinetes, 57 heterocyst-specific proteins (33 novel proteins), including nif gene products, and 485 proteins exclusively in vegetative cells. HAVe model was proposed that akinetes, unlike the typical spores of bacteria, perform unique biochemical functions that collaborate with both heterocysts and vegetative cells. Regardless of nitrate availability, some vegetative cells of Anabaena cylindrica are programed to differentiate semi-regularly spaced, single heterocysts along filaments. Since heterocysts are non-dividing cells, with the sole known function for solar-powered N2-fixation, is it necessary for heterocyst to retain entire genome (7.1Mb) from its progenitor vegetative cell? By sequencing the genome of isolated heterocyst, I discovered that at least six DNA elements (0.12 Mbp) are deleted from the heterocyst genome during heterocyst development. The six-element deletions restore five genes (nifH1, nifD, hupL, primase P4, acyl_5725 (hypothetical protein) that were interrupted in the genome of vegetative cells. More deletions are expected to be identified in the completed genome of heterocyst, a uniquely solar-powered, oxic N2-fixing cell. To my best knowledge, this is the first report that (1) different genomes may occur in distinct cell types in a single bacterium; and (2) genome editing is coupled to cellular differentiation and/or cellular function in a multicellular cyanobacterium. In response to environmental changes, Anabaena cylindrica differentiate three cell types, vegetative cells for photosynthesis, heterocysts for nitrogen fixation, and akinetes for stress survival. Cell-surface polysaccharides play important roles in bacterial ecophysiology. In this study, specific cell-surface sugars were discovered in heterocysts, akinetes and vegetative cells of A. cylindrica using 20 fluorescein labeled lectins. Both Nacetylglucosamineor- binding lectins WGA and succinylated WGA bound specifically to the vegetative cells. Akinetes bound to three mannose-binding lectins (LCA, PSA, and ConA), and one of the galactose-binding lectins (GSL-I). ConA also bound to heterocyst, and the binding was diminished in the heterocysts of an all4388 mutant, in which the putative polysaccharide export protein gene all4388 was disrupted. Above proteomics, genomics and genetic research greatly added to our understanding of the cell development in A. cylindrica and clarify the patterns of gene expression in heterocysts, akinetes and vegetative cells, which pave the way for further study of Anabaena cylindrica.

Library of Congress Subject Headings

Cyanobacteria.
Anabaena.
Nitrogen-fixing microorganisms.
Proteomics.

Description

Includes bibliographical references (pages 106-138)

Format

application/pdf

Number of Pages

179

Publisher

South Dakota State University

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Rights Statement

In Copyright