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- In vivo analysis of Arabidopsis FtsZ isoforms
- Schmitz, Aaron James
- Electronic Theses & Dissertations
Chloroplasts are organelles derived from the ancient endosymbiosis between a cyanobacterium and a primitive eukaryote. These organelles are essential for algae and plants for their many functions, including photosynthesis, biosynthesis of a wide array of essential molecules, and reduction of sulfur and nitrogen. Chloroplasts cannot form de novo and their numbers are maintained through the process of binary fission. This division process requires several genes originally encoding bacterial...
Show moreChloroplasts are organelles derived from the ancient endosymbiosis between a cyanobacterium and a primitive eukaryote. These organelles are essential for algae and plants for their many functions, including photosynthesis, biosynthesis of a wide array of essential molecules, and reduction of sulfur and nitrogen. Chloroplasts cannot form de novo and their numbers are maintained through the process of binary fission. This division process requires several genes originally encoding bacterial cell division factors. FtsZ is one of these genes. FtsZ is present in most bacteria and encodes a cytoskeletal protein that is structurally similar to tubulin. FtsZ polymerizes into a ring structure (Z-ring) at mid-cell prior to cell division in bacteria and also forms a Z-ring within the stroma of chloroplasts and other plastids types in plants. However, in the green lineage FtsZ has split into two phylogenetically-distinct families called FtsZ1 and FtsZ2. Both families have been shown to colocalize to the Z-ring and interact with themselves and each other. Chloroplast division, like cell division in bacteria, is sensitive to small decreases or increases in FtsZ protein levels, which result in division defects and fewer, enlarged chloroplasts. Therefore, distinguishing the relationship between the encoded FtsZ protein isoforms based upon ftsZ null or overexpression mutants is not feasible. The focus of this work was to resolve the functional relationship and distinguishing features of FtsZ isoforms in Arabidopsis - our chloroplast division model. Stable transformation of Arabidopsis ftsZ mutants followed by careful examination of complemented chloroplast division defects and FtsZ protein levels was the predominant approach for these studies.FtsZ2-1 complemented chloroplast division defects of plants lacking FtsZ2-2, and vice versa, near the previously quantified protein levels expected for complete FtsZ2 substitution. Therefore, I conclude that the two AtFtsZ2 isoforms are functionally redundant. Subsequently, I determined that FtsZ1 cannot substitute for FtsZ2 protein, and vice versa, since chloroplast division defects remained. In a related study, though both FtsZ1 and FtsZ2 are required for maintenance of chloroplast numbers, the generation of fully viable Arabidopsis plants lacking FtsZ, yet maintaining one chloroplast per cell, indicated that an FtsZ-independent mode of chloroplast partitioning exists in higher plants.FtsZ1 and FtsZ2 proteins diverge significantly at their C-termini where only the FtsZ2 family has a conserved motif found in bacteria. This motif is critical for the interaction with Z-ring promoting factors in bacteria and with ARC6 in plants. By swapping the C-termini and substituting the resulting chimeric FtsZ proteins in vivo, I demonstrate that neither C-terminus fully defines the unique functions of FtsZ1 or FtsZ2. Though I also show that the C-termini are required for the full function of each FtsZ family, these results indicate that other regions contribute significantly to FtsZ function. Related experiments also indicate that ARC3, a negative regulator of Z-ring formation, interacts with FtsZ2 in addition to FtsZ1. Together these results have clarified FtsZ functional relationships and laid significant groundwork for future analyses of FtsZ and their regulators.