Sizes of (plastid) cpDNA

1. Sizes of (plastid) cpDNA • Range is 70,000 bp (70 kb) to ~2,000,000 bp (2,000 kb), but most are less than 250,000 bp (250 kb) • Land plants typically 120 – 170 kb (70 kb – Epifagus; ~2,000 kb – Acetabularia)

2. Sizes of cpDNAs from diverse plants From Kloppstech, Westhof et al.

3. Parasitic plant, no photosyn. Epifagus virginiana -beechdrops From U. Wisconsin-Madison Botany Dept.

4. Organization of typical (angiosperm) chloroplast chromosome • inverted repeats (IRa and IRb) separate circle into large and small single-copy regions (LSC and SSC, respectively) • IRs always contain the rRNA (rrn) genes, but also contain other genes • ~125 genes are found, encoded on both strands, without much overlap

5. Tobacco cpDNA (Sugiura lab) From Kloppstech, Westhof et al.

6. cpDNA Gene Content Most cp genes fall into 2 functional groups: • genes involved in the genetic apparatus (replication, transcription,translation) • genes involved in photosynthesis Also genes for protein degradation, fatty acid synthesis, and respiration (chlororespiration?).

7. Gene identification, or "Sorting out Gene-Protein Relationships" Two basic approaches: protein --> DNA DNA  protein

8. Gene nomenclature Based on bacterial naming system, which uses lower case letters, and a descriptive prefix, based on the probable function. If the gene product is part of a multi-subunit complex, a letter of the alphabet is used to denote different subunits. Examples: psa for genes of photosystem I (psaA, psaB, etc.) psb for genes of photosystem II (psbA, psbB, etc.) A non-conforming example: rbc for genes encoding ribulose-1,5-bisphosphate carboxylase (RuBPCase) - RuBPCase has two subunits, large and small - the genes are rbcL and rbcS; rbcL is in cpDNA, rbcS is encoded in the nucleus

9. accD

10. Comparative organization of cpDNA among land plants and green algae • The length of the IR regions vary in different plant families. • There is no IR in certain plants (e.g., legumes). • Significant differences in gene order between distantly related species, but relatively minor differences in gene content (e.g., between land plants and Chlamydomonas).

11. Cp Genome in non-green algae In evolutionarily ancient (or distant) algae, such as reds (rhodophytes) or chromophytes (Chl a/c-containing brown or golden algae) the cp genome can be quite different: • contains more genes (up to 2x more, 250), many of which are in the nucleus in green plants • sometimes have multiple large circles Dinoflagellates have very weird Cp genome made up of many small gene-sized plasmids.

12. Chrysophytes - type of Chromophyte Ceratium “Dinos” Dinophysis Porphyra A Rhodophyte

13. Chloroplast Origins & Evolution • The plastid genome is fairly conserved in evolution (compared to nuclear or mito.). • It originated from the endosymbiotic associations that formed eukaryotic cells "Endosymbiotic Hypothesis“. • The precursor endosymbiont was a cyanobacterial-like organism. • Most of the endosymbiont’s genes were either lost, or transferred to the nucleus early in evolution.

14. Can we find instances of more recent gene transfer from plastid to nucleus? 1. tufA gene (chloroplast translation elongation factor Tu) is in cpDNA of most green algae, but in the nucleus in land plants. 2. rpl22 gene (chloroplast ribosomal protein) in cpDNA in all plants except legumes, where it’s (only) in the nucleus. Analysis of this gene suggests it was in the nucleus a long time before the chloroplast gene was lost. “phylogenetic analysis shows that rpl22 was transferred to the nucleus in a common ancestor of all flowering plants, >100 million years before it was lost from the legume chloroplast lineage” Conclusion: Gene transfer to nucleus still going on, and some genes are more likely to transfer than others.

15. Phylogenetic evidence suggests a common origin for all plastid genomes.However, some chloroplasts were acquired secondarily. Chromophytes, dinoflagellates and euglenoids have 3 (and sometimes 4) membranes around the chloroplast.

16. Euglenoids have 3 membranes around chloroplast: - outer & inner envelope membranes - extra membrane resembling an ER membrane - also have many “animal” characters It is suggested that a photosynthetic eukaryote (green alga) was the endosymbiont, and its chloroplast was retained.

17. Chloroplast ER (CER) with 2 membranes, making 4 around this organelle in the chromophyte, Olisthodiscus S. Gibbs

18. In cryptomonads andchlorarachniophytes, there is even a remnant of the endosymbiont’s nucleus, called the Nucleomorph. In cryptomonas, itis made up of 3 small chromosomes (~600 kb) with 510 genes, ~30 for plastid proteins. Also has genes for gene expression. Ref: Douglas et al. (2001) Nature 410:1091

19. Cryptomonadcell w/host (blue) & endosymbiont parts (red) http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/E/Endosymbiosis.html

20. Keeling, 2004, Am. J. Bot. 9:1481

21. Primary Endosymbiosis Secondary Endosymbiosis Tertiary or Serial Endosymbiosis Keeling, 2004, Am. J. Bot. 9:1481

22. Nucleomorph genes (Green lineage) • P.R. Gilson, V. Su, C. H. Slamovits, M.E. Reith, P.J. Keeling, and G. I. McFadden (2006) Complete nucleotide sequence of the chlorarachniophyte nucleomorph: Nature’s smallest nucleus. Proc. Natl. Acad. Sci. USA 103: 9566-9571. • 331 genes on 3 chromosomes ( ~373,000 bp) • 17 genes for plastid proteins • tiny introns (20 nt) (GT….AG)

23. Elysia chlorotica Sea slug with active chloroplasts from a heterokont alga (Vaucheria). Chloroplasts stay active for at least 8 months. “Kleptoplasty” – growing with stolen plastids Rumpho, M.E., Summer, E.J. & Manhart, J.R. (2000) Solar-Powered Sea Slugs. Mollusc/Algal Chloroplast Symbiosis. Plant Physiology, 123: 29-38.

24. Vaucheria Heterokontophyta (Xanthophyceae) A few nuclei (rounder) and many chloroplasts in these giant cells. Plastids acquired secondarily, are of “red algal origin”.

25. Isolation of Functional Chloroplasts from the Sacoglossan Mollusc Elysia viridis Montague; M.L. Williams; A. H. Cobb. New Phytologist, Vol. 113, pp. 153-160 (1989) Codium green & marine

26. How do chloroplasts remain active for several months? • 2 possibilities: • They are unusual and encode many genes that are found in the nucleus in other plants. • - No, but did lack psbO gene • The slug (Elysia) has acquired genes for plastid proteins in its nucleus and provides the proteins to the stolen chloroplasts. • - Found psbO in the nucleus of Elsyia and Vaucheria. • If 2 is correct, then there should be many others!! Rumpho et al. (2008) Proc Nat l Acad Sci 105, 17867

27. Endosymbiosis has played a major role in the evolution of life on earth, and will likely continue to do so. Maybe animals, or animal-plant hybrids that photosynthesize are out there? Or have come and gone (extinct)?