Mechthild Pohlschroder

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Graduate Group Affiliations

Contact information
201 Leidy Labs
415 University Avenue
Philadelphia, PA 19104-6018
Office: 215 573-2283
Fax: 215 898-8780
Vordiplom (Biology)
University of Muenster, Germany, 1989.
PhD (Microbiology)
University of Massachusetts, Amherst, 1994.
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Description of Research Expertise

Research Interests
Prokaryotic Protein Translocation across Hydrophobic Membranes and Their Substrates.

Key words: archaea, protein secretion, porkaryotes, haloarchaea, type IV pili, flagella, biofilm.

Description of Research
Description of Research

To remain viable, an organism must translocate a subset of its proteins into or through hydrophobic cellular membranes. The proteins translocated through these membranes play a variety of invaluable functional role in critical cellular processes, which include nutrient acquisition, toxin secretion, signal transduction and the formation of extracytoplasmic protein complexes, such as pili and flagella.

The pathways used by cells to facilitate protein translocation include the Sec system, which transports proteins in an unfolded conformation and is thought to be the major translocation pathway used by most organisms, and the twin-arginine translocation (Tat) system, which transports unfolded proteins. These translocation pathways have been characterized in bacteria and in eukaryotes, and have recently been described in archaea.

Our lab focuses on identifying and characterizing the molecular components in these translocation pathways in archaea. Characterization of these pathways in archaea will allow us to: 1) define general principles for each type of translocation pathway; 2) determine the bases for the preferential transport of specific proteins via the Sec or Tat pathway; and 3) based on the similarities and differences the pathways in archaea, bacteria and eukaryotes, determine evolutionary relationships.

We study these translocation pathways in the model system Hfx. volcanii, a haloarcheon that is amenable to genetic and biochemical manipulation and analyses. In addition to addressing the aims stated above, by elucidating the biological processes that are dependent on the substrates translocated by these pathways in haloarchaea, which thrive in salt conditions approaching saturation, we may be able to identify specific adaptations made by this species to high salt concentrations. We are complementing our in vivo studies of the mechanisms underlying protein translocation with computational in silico analyses of pathway components and the substrates of these pathways.

The haloarchaeal Sec pathway: Most proteins pass through the endoplasmic reticular membranes of eukaryotes and the cytoplasmic membranes of bacteria via a proteinaceous pore known as the Sec translocon. While the core components of these pores are evolutionarily conserved in bacteria and eukaryotes, the specific functions of most of these components are not well understood. Interestingly, the archaeal Sec pathway contains a combination of bacterial and eukaryotic Sec component homologs. Curiously, no archaeal homolog of a translocation ATPase has yet been identified. Using a combination of genetic, biochemical and in silico approaches, we have been identifying and characterizing Hfx. volcanii homologs of known Sec components and we are also attempting to identify as yet unknown Sec pathway components, including an archaeal Sec translocation ATPase.

The haloarchaeal Tat pathway: Many archaeal species possess unique adaptations that allow survival in extreme environments. In haloarchaea, in silico and in vivo evidence obtained by our lab suggests that, while the Sec pathway is crucial for haloarchaeal growth, haloarchaea route most secreted proteins through the Tat pathway, pohlschr/, which may be an adaptation to the high salt concentrations found in their natural environments. This unique characteristic makes studying the haloarchaeal Tat pathway particularly useful since a wide range of substrates is available for use in characterizing the functions of known Tat components as well as for screens and selections that can be used to identify additional Tat components. Moreover, revealing the significance of haloarchaea-specific Tat pathway characteristics and comparing these characteristics to those of non-haloarchaeal pathways may provide important information concerning the efficient use of the Tat pathway and lead to a better understanding of pathway mechanisms in general. Using this system, we can also address crucial problems such as: 1) determining the dynamics of pore assembly; 2) discovering how proteins having drastically differing sizes can be translocated while maintaining the essential semipermeability of the membrane; and 3) identifying the selective pressures that target substrates to the Tat or Sec pathway.

Finally, it should be noted that eukaryotic Tat pathway components have only been identified in chloroplasts. Since Tat mutants of pathogenic Escherichia coli and Pseudomonas aeroginosa, along with Tat mutants of other bacterial pathogens, are attenuated for virulence, certain components of this pathway may be attractive drug targets.

Archaeal type IV pili: Although bacterial flagella and archaeal flagella have similar functions, that is, they are responsible for swimming motility in these prokaryotes, the synthesis and structures of these protein complexes are very different. In fact, the synthesis of archaeal flagella closely resembles that of bacterial type IV pili and the structural subunits of these complexes, which are secreted via the Sec pathway, are also related. Since Hfx. volcanii are capable of biological processes that in bacteria require the involvement of type IV pili, based on sequence data generated for archaeal flagellin, we developed a software program that allowed us to identify several operons in the Hfx. volcanii genome that encode pilin-like proteins, as well as similar operons in many other archaeal genomes. We have now determined the temporal expression patterns for these Hfx. volcanii operons and we are currently optimizing conditions for their expression. We have also generated several mutant strains and we are attempting to determine the roles these pilin-like proteins play in surface adhesion, autoaggregation, twitching motility and conjugation, all processes that in bacteria require functional type IV pili.

Rotation Projects for 2006-2007
1. Characterization of H. volcanii homologs of bacterial and eukaryotic Sec or Tat components with respect to their function in archaeal protein translocation. These studies may include analyses of their interactions with other translocation components or in vivo analyses of knockout-or conditional mutants of the cloned genes.

2. Genetic screens and selections to identify components that are involved in the Tat or Sec secretion pathways. Approaches will be used that have successfully been applied in the bacterial and eukaryotic systems. These studies may allow us to identify components that are archaea specific but may also reveal components that have previously not been identified in bacteria and eukaryotes.

3. Characterization of Hfx. volcanii type IV pilus-like structures: We have identified 6 operons containing genes that encode putative pilins. The distinct sequences and expression profiles of these pilins suggest that the pili are involved in different functions. The project would involve the biochemical characterization of the subunits as well as making knockouts of some of these genes and determining their phenotypes.

Lab personnel:
Kristin Toscano, Graduate Student
Manuela Tripepi, Graduate Student
Ameya Nanivadekar, Undergraduate Student
Jason You, Research Associate

Selected Publications

Albers S. and M. Pohlschroder: Archaeal Type IV pili. Extremophiles in press, 2009.

Szabo, S. A. Oliveira Stahl, Z. Albers, A. Driessen, J. Kissinger and M. Pohlschröder: Identification of diverse archaeal proteins with class III signal peptides cleaved by distinct archaeal prepilin peptidases. J. Bacteriol. 189: 772-778, 2007.

Widdick, D., K. Dilks, G. Chandra, A. Bottrill, M. Naldrett, M. Pohlschröder and Tracy Palmer: Secretome analysis reveals that the twin-agrinine translocation (Tat) pathway is a major route of protein export in Streptomyces coelicolor. PNAS 106: 17927-17932, 2007.

Dilks, K. M. I. Gimenez and M. Pohlschröder: Haloferax volcanii twin arginine translocation substrates include soluble secreted, C-terminally anchored and lipoproteins. Mol. Microbiol. 66: 1597- 1602, 2007.

Pohlschröder, M., E. Hartmann, M, N. J. Hand, K. Dilks, M. A. Haddad: Diversity and evolution of protein translocation. Invited review, Annual Reviews in Microbiology 59: 91-111, 2005.

Haddad, A., R. W. Rose, M. Pohlschröder: The Haloferax volcanii FtsY homolog is critical for haloarcheal growth but does not require the A-domain. J. Bacteriol. 187: 4015-4022, 2005.

Dilks, K. M. I. Gimenez, and M. Pohlschröder: Genetic and biochemical analysis of the twin-arginine translocation pathway in halophilic archaea. J. Bacteriol. 187: in press, 2005.

Dilks, K., R.W. Rose, Enno Hartmann, and M. Pohlschröder: Prokaryotic use of the twin arginine translocation pathway: A Genomic Survey. J. Bact. 185: 1478-83, 2003.

Adaptation of proteins secretion to extremely high salt concentrations by extensive use of the twin arginine translocation pathway: Rose, R.W., T. Brüser, J.C. Kissinger, and M.Pohlschröder. Mol. Microbiol. 45: 943-50, 2002.

Hand, N. J., A. Laskewitz, R. Klein, and M. Pohlschröder: Archaeal and bacterial SecD and SecF homologs exhibit striking structural and functional conservation. J. Bacteriol. 188: 1251-1259, 2006.

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Last updated: 08/21/2009
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