Chloroplast DNA is usually circular with a long single copy section (LSC) and short single copy section (SSC) separated by inverted repeats. It contains around 100-200 genes encoding proteins, tRNAs, and rRNAs. These genes are involved in chloroplast gene expression and photosynthesis. Most proteins are nuclear gene products imported into chloroplasts, but the large subunit of RuBisCO is encoded by a chloroplast gene. Chloroplast genes are arranged in operon-like units and co-transcribed using different promoters. There are two RNA polymerases - PEP and NEP - that transcribe chloroplast genes. PEP resembles the bacterial polymerase while NEP resembles phage polymerases.

1. Gene Expression of Chloroplast
Chloroplast DNAs are usually a circular double stranded DNA molecules containing a long
single copy section (LSC) and a short single copy section (SSC) separated by two inverted
repeats. Normally chloroplast genome (cp genome) contains about 100- 200 base pairs and codes
for about 60 proteins 30tRNAs and 4rRNA in a higher plant. Some of these genes encode
components such as RNA polymerase, ribosomal proteins, tRNAs, and rRNAs which are
involved in chloroplast gene expression machinery and subunits of photosynthetic enzymes such
as Rubisco, PSII, the cytochrome b6f complex, PSI, ATP synthase and NADH dehydrogenase.
Some genes encode proteins like ClpP, AccD, Ycf1, and Ycf2 which are involved in other
metabolic processes. Most of the proteins with in chloroplast are nuclear gene products that are
synthesized in the cytosol and , imported and some are synthesized within chloroplast. RuBisCO
(Ribulose-1, 5-bisphosphate carboxylase/oxygenase) the major enzyme involved in carbon
fixation process and photo respiration within chloroplast. Nuclear gene (rbcS) encodes the small
subunit, whereas a single gene (rbcL) located on multiple copies of DNA in the chloroplast
encodes the large subunit. Aassembly is occurred correctly in the presence special folding
proteins called chaperonins.Most of the cp genes are arranged in operon-like units and co-
transcribed. For examples psbD-psbC gene cluster is an operon and in psbD-psbC gene
transcription, a different (closer) promoter is used in the presence of the light called light-
responsive promoter (LRP). Nearly there are 30 promoters in higher plant cp DNA and a
"transcription unit" is determined by the position of the promoter (5') and terminator (3') signals.
Most of the promoters resembles the major E. coli σ70 (or -10,-35) promoter. In some promoters
the distance between -10 and -35 regions is critical. For sofme genes, no -10 and -35 regions as
they have other promoter type. The distance between -10 and start point (+1) is less critical. The
consensus sequences at -10,-35 and +1 are highly variable among species. Terminators not
clearly defined, but tRNA genes seem to be transcription terminators. Transcription regulation is
done mainly by the regulation of posttranscriptional steps and also at initiation step using
alternative promoters. Transcription rate is affected due to the presence of some genes (psbD) by
upstream sequences that bind regulatory proteins and the “promoter strength”.There are 2 major
CP RNA polymerases in vascular plants. They are E. coli or eubacterial-like polymerase (PEP,
plastid-encoded polymerase) and phage-like polymerase (NEP, nuclear-encoded polymerase).
PEP is composed of core and Sigma factor. Core is made up of 4 subunits, α2 ββ' (α is encoded
by the rpoA gene, β is encoded by the rpoB gene, β' is encoded by the rpoC1 and rpoC2 genes).
Sigma factor is needed to initiate transcription as it recognizes -10,-35 regions for some
promoters. This polymerase is nhibited by rifampicin. NEP polymerase consisted of Catalytic
subunit is similar to the 1-subunit phage (e.g., T7) and mitochondrial RNA polymerases and it is
coded by a nuclear gene. It identifies the promoter consisted of a single region of 7-10 bp. It is
insensitive to rifampicin. NEP is more important early in plastid development, when plastid
transcription and translation is relatively low. This transcribes rRNA, rpo and other genetic
functions genes (GFG).PEP is more important in mature chloroplasts as it transcribes some GFG
genes and most of photosynthesis genes.CP pre-mRNA is processed by cleavage(s) or splicing or
both. CP mRNAs are not polyadenylated, and are not capped. Nucleolytic Cleavages are
occurred in 2 ways. Endonucleases cut internal regions specifically (e.g. in between genes) and
exonucleases trim at 3' or 5'-ends, processively less specifically. Inverted repeats (IR) occur at 3'-
end of most cp protein encoding genes also they act as processing sites, determining the 3'-end of
mRNAs. Proteins recognize the 3'-IR, bind and stop the processive exonucleases. An

2. endonuclease cleaves at the 3’-IR. Translation machinery is similar to bacteria. Ribosomes are
70S (composed of L (50S) and S (30S) subunits) and contain 23S (L), 16S (S), and 5S (L)
rRNAs. Each subunit (L and S) contains about 30 proteins. Initiation factors involved are if1, if2,
if3 and longation factors involved are ef-Tu, ef-Ts, and G. Translation is initiated with fmet
(formylated Met). As many cp mRNAs contain a Shine-Dalgarno sequence preceding the first
codon; it base-pairs with the 3'-end of 16S rRNA and translation is proceeded.In some cases
Shine-Dalgarno sequence suppress the translation and proteins that bind the 5’ UTRs of mRNAs
promote translation. Start codon is very important for starting translation at right codon. Internal
ORFs are translated in to a polycistronic transcript.
Reference:-
Barkan A. (2011). Expression of Plastid Genes: Organelle-Specific Elaborations on a
Prokaryotic Scaffold. Plant Physiology. 155: 1520–1532.
Buchanan, B. B. Gruissem, W. Jones, L.R. 2005. Biochemistry & molecular biology of plants,
6th edition. Courier Companies, Inc., U.S.A, pp. 508- 565.
P.R. Archie. (2001). Rubisco. Encyclopedia Of Life sciences. 6: 125–149.
http://slideplayer.com/slide/6186650/.htm (2015.09.22).