Structure and function of the plant plastid-encoded RNA polymerase

Vergara Cruces, Ángel (2025) Structure and function of the plant plastid-encoded RNA polymerase. Doctoral thesis, University of East Anglia.

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Abstract

In plants, photosynthesis happens in specialized organelles called chloroplasts. Chloroplasts derive from endosymbiosis of a cyanobacterial ancestor and because of this, they possess their own genome and gene expression machinery, which resemble those of bacteria. Chloroplast gene expression is critical for the regulation of photosynthesis, since central components of photosynthetic complexes are encoded in the chloroplast genome. Expression of photosynthetic genes is dependent on the plastid-encoded RNA polymerase (PEP). PEP is a 1 MDa complex of a bacterial-type multi-subunit RNA polymerase, and at least 12 additional PEP-associated proteins (PAPs). Although PAPs are not homologous to transcription factors of bacteria, they are essential for expression of photosynthetic genes. Loss of function of most PAPs result in plants that are unable to develop chloroplasts and perform photosynthesis. Despite their importance, the molecular roles of PAPs in chloroplast transcription are not fully understood.

The subject of this thesis is the characterization of chloroplast transcription at the molecular level. PEP has not been previously obtained in sufficient purity for rigorous biochemical and structural characterization. Here, I present the native purification of PEP from Sinapis alba. Proteomic analysis of purified PEP provided a definitive composition of the stable PEP complex. Structural models of the PEP complex and a PEP transcription elongation complex were obtained by cryogenic electron microscopy (cryo-EM). These models revealed how the PAP proteins interact with the core polymerase and with DNA and nascent mRNA. In addition, cryo-EM analysis of a PEP complex with the conserved transcription elongation factor NusG is also presented. This revealed novel modes of interaction of chloroplast NusG with PEP that have not been observed in previously characterized bacterial transcription complexes. This work lays the foundation for a more detailed understanding of the regulation of chloroplast gene expression and its role in plant development and adaptation.

Item Type:	Thesis (Doctoral)
Faculty \ School:	Faculty of Science > School of Biological Sciences
Depositing User:	Chris White
Date Deposited:	21 May 2026 09:49
Last Modified:	21 May 2026 09:49
URI:	https://ueaeprints.uea.ac.uk/id/eprint/103123
DOI:

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