The Importance of CO2 Recapture in the CO2 Concentrating Mechanism of Chlamydomonas Reinhardtii

Author

Ashwani Rai, Louisiana State University and Agricultural and Mechanical CollegeFollow

Degree

Doctor of Philosophy (PhD)

Department

BIOLOGICAL SCIENCES

Document Type

Dissertation

Abstract

The aim of this thesis is to investigate the CO₂ concentrating mechanism (CCM) of Chlamydomonas reinhardtii and to develop a quick method for estimating the activity of carbonic anhydrases (CAs). The first project demonstrates that there are two almost identical mitochondrial CAs in C. reinhardtii, CAH4 and CAH5, that help to maintain photosynthesis and minimize the leak of CO₂ generated by respiration and photorespiration. We used an RNAi approach to reduce the expression of CAH4 and CAH5 so that their physiological functions could be studied. RNAi mutants with low expression of CAH4 and CAH5 have impaired rates of photosynthesis under ambient air levels of CO₂ (0.04% [v/v] CO₂ in air). These strains are not able to grow in very low-CO₂ conditions (2 in air) but grew normally on elevated CO₂ (5% [v/v] CO₂ in air)_. These RNAi mutants had a poor affinity for inorganic carbon (C_i = CO₂ + HCO₃^-) and their ability to accumulate C_i is impaired. We hypothesize that CAH4 and CAH5 work in the mitochondria to convert the CO₂ released from respiration and photorespiration and the CO₂ leaked from the chloroplast to HCO₃^-, thus “recapturing” this potentially lost CO₂.

In the second project, we developed a method to estimate CA activity using a mutant complementation approach in Saccharomyces cerevisiae (hereafter referred to as yeast) in order to study the role of CAs in photosynthetic organisms. In studies based on this method, we found that CAH4/5 and CAH3 from C. reinhardtii, βCA3 from Arabidopsis thaliana and different modified forms of Human CA II are all active CAs. We also determined that codon optimization can improve protein expression in the heterologous yeast system. It is important to know if CAs are active, as introducing CA activity in the correct location in a plant chloroplast could accelerate efforts to improve crop plant photosynthetic efficiency. It will also help to understand how CAs function across different locations in photosynthetic organisms.

Date

7-5-2022

Recommended Citation

Rai, Ashwani, "The Importance of CO2 Recapture in the CO2 Concentrating Mechanism of Chlamydomonas Reinhardtii" (2022). LSU Doctoral Dissertations. 5913.
https://repository.lsu.edu/gradschool_dissertations/5913

Committee Chair

Moroney, James V.

DOI

10.31390/gradschool_dissertations.5913