The Engineered Chloroplast Genome Just Got Smarter

Abstract

Chloroplasts are known to sustain life on earth by providing food, fuel and oxygen through the process of photosynthesis. However, the chloroplast genome has also been smartly engineered to confer valuable agronomic traits and/or serve as bioreactors for production of industrial enzymes, biopharmaceuticals, bio-products or vaccines. The recent breakthrough in hyper-expression of biopharmaceuticals in edible leaves has facilitated the advancement to clinical studies by major pharmaceutical companies. This review critically evaluates progress in developing new tools to enhance or simplify expression of targeted genes in chloroplasts. These tools hold the promise to further the development of novel fuels and products, enhance the photosynthetic process, and increase our understanding of retrograde signaling and cellular processes.

Codon Optimization to Enhance Expression Yields Insights into Chloroplast Translation

“…Transcript abundance is attributed to the high copy number of transgenes and the strength of the psbA promoter. Among more than 150 transgenes expressed in chloroplasts, more than 75% utilized psbA regulatory sequences (Jin and Daniell, 2015;Daniell et al, 2016aDaniell et al, , 2016b. In addition, three ribosome-binding regions in the 5ʹ UTR of psbA recruit ribosomes and efficiently form the translational initiation complex (Zou et al, 2003).…”

“…The 5ʹ UTR of psbA also showed the highest translation activity in vitro among 11 5ʹ UTRs investigated (Yukawa et al, 2007). Therefore, among 140 transgenes expressed in chloroplasts, more than 75% use the psbA regulatory sequences (Jin and Daniell, 2015;Daniell et al, 2016aDaniell et al, , 2016b. Most importantly, compatibility between the 5ʹ UTR of psbA and its coding region is important for efficient translation initiation (Nakamura et al, 2016).…”

“…Among more than 115 transgenes expressed via the chloroplast genome, 84 use the psbA regulatory sequence (Jin and Daniell, 2015;Daniell et al, 2016aDaniell et al, , 2016b. A recent study (Nakamura et al, 2016) shows the absence of any detectable translation when codons for the tat coding sequence of HIV-1 were optimized using all 79 tobacco chloroplast mRNAs and regulated by the psbA 5ʹ UTR (Nakamura et al, 2016), but the same sequence was expressed well using the phage T7 GENE10 5ʹ UTR.…”

“…A wide range of proteins, from very small antimicrobial peptides (Lee et al, 2011) or hormones (Boyhan and Daniell, 2011;Kwon et al, 2013) to very large proteins encoded by bacterial, viral, fungal, animal, and human genes, have been expressed successfully in plant chloroplasts (DeGray et al, 2001;Daniell et al, 2009;Verma et al, 2010;Shenoy et al, 2014;Sherman et al, 2014;Shil et al, 2014). Most importantly, expressed proteins are highly stable when lyophilized plant cells are stored at ambient temperature (Kwon et al, 2013;Lakshmi et al, 2013;Kohli et al, 2014;Jin and Daniell, 2015). Therefore, oral delivery of proinsulin or exendin-4 reduced blood sugar levels similar to injected proteins (Boyhan and Daniell, 2011;Kwon et al, 2013).…”

Rapid production of therapeutic proteins using plant system

“…Using this approach a whole range of therapeutic proteins has been produced in the plant tissue. Depending upon the use of a specific promoter, expression can be achieved uniformly throughout the plant or it can be limited up to the expression in plant seed, organs and sub-cellular organelles [28][29][30][31][32][33] . Apart from this, the focus of future research is in the production of oral vaccines in the edible plants/ fruits, such as tomatoes and bananas.…”