Research Summary

The chloroplast and mitochondrion are the two energy-producing organelles in eucaryotic cells. Both organelles contain their own DNA, but most of the proteins required for their biogenesis are encoded by the nuclear genome and synthesized in the cytosol. The chloroplast carries out the unique reactions of photosynthesis, and also houses an amazing array of biosynthetic pathways required for plant development that depend on nuclear gene expression and selective targeting of precursor proteins to the correct organelle. The major goal of our research is to understand the pathway of protein import into the chloroplast, and elucidate at the molecular level the key components involved and how they carry out their functions. Our studies identified a general stromal processing peptidase, called SPP, that removes targeting signals from nearly all proteins that enter the chloroplast, releasing them to attain their mature conformations and assemble into active multisubunit complexes. Our biochemical and transgenic plant studies have demonstrated that SPP is essential for plant survival, and its structure is highly conserved. We are investigating how SPP selectively recognizes its unique substrates, and the mechanisms underlying its activity. To broaden our understanding of the chloroplast import pathway, we have developed a novel genetic screen in the model plant Arabidopsis that depends on two independent reporters: green fluorescent protein (GFP) and an herbicide resistant enzyme that is located only in chloroplasts. A class of chloroplast import deficient mutants has been identified where GFP is mislocalized to the nucleus. A genetic analysis has the potential to identify:

1) new components required for protein sorting in the cytosol and transport across the chloroplast membranes,

2) distinct import pathways employed by the large diversity of precursors targeted to the organelle in different tissues,

3) genes that regulate the expression and the assembly of the import machinery, and

4) developmental and physiological factors that influence import specificity and efficiency.

Our results should contribute to an understanding of protein sorting and trafficking in a eucaryotic cell that have evolved to create its many organelles with specialized functions.

Selected Publications

Ponpuak, M., Klemba, M., Park, M., Gluzman, I., Lamppa, G. and Goldberg, D. 2007. A role for falcilysin in transit peptide degradation in the Plasmodium falciparum apicoplast. Molecular Microbiol.: 63: 314-334. (PubMed)

Richter, S., Zhong, R. and Lamppa, G. (2005) Function of the stromal processing peptidase in the chloroplast import pathway. (Review) Physiol. Plant. 123: 362-368.

Rudhe, C., Clifton, R., Chew, O., Zeman, K., Richter, R., Lamppa, G., Whelan, J., and
Glaser, E. 2004. Processing of the dual targeted precursor protein of glutathione reductase in mitochondria and chloroplasts. J. Mol. Biol. 343: 639-647. (PubMed)

Jin, R., Richter, S., Zhong, R. and Lamppa, G. K. (2003). "Expression and import of an active cellulase from a thermophilic bacterium into the chloroplast both in vitro and in vivo." Plant Mol Biol 51: 493-507. (PubMed)

Zhong, R., Wan, J., Jin, R. and Lamppa, G. (2003). "A pea antisense gene for the chloroplast stromal processing peptidase yields seedling lethals in Arabidopsis: survivors show defective GFP import in vivo." Plant J 34: 802-12. (PubMed)

Richter, S. and Lamppa, G. K. (2003). "Structural properties of the chloroplast stromal processing peptidase required for its function in transit peptide removal." J Biol Chem 278: 39497-502. (PubMed)

Richter, S. and Lamppa, G. K. (2002). "Determinants for removal and degradation of transit peptides of chloroplast precursor proteins." J Biol Chem 277: 43888-94. (PubMed)

Richter, S. and Lamppa, G. K. (1999). "Stromal processing peptidase binds transit peptides and initiates their ATP-dependent turnover in chloroplasts." J Cell Biol 147: 33-44. (PubMed)