Robert Marsh

Senior Lecturer III - Biological Sciences
robert.marsh@utdallas.edu
972-883-2511
FN2202
Tags: Biology

Professional Preparation

Ph.D. - Molecular Biology
Vanderbilt University - 1971
B.S. - Chemistry and Mathematics
Western Kentucky University - 1965

Research Areas

Research and Expertise

Overview

Dr. Marsh's research interests focus on subcellular structure. He is currently involved in projects concerning the identification and characterization of nuclear matrix proteins, cell surface lectins, and the protein cross-linking enzyme transglutaminase.

Physarum Transglutaminase and its Substrates

Physarum polycephalum is a lower eukaryote that can grow as plasmodia containing multiple nuclei.  Chemical damage of the plasmodia activates a Ca2+-dependent transglutaminase that catalyzes cross-linking of proteins in nuclei and cytoplasm by acyl group transfer between glutamine side chains and the e-amino group of lysine residues. Extraction of the plasmodia with SDS leaves filamentous envelopes filled with residual, ghost nuclei (Figure 1) that are protease sensitive and nuclease resistant.  Protein cross-linking may block the spread of damage throughout plasmodia and prevent loss of cytoplasm.   

We have identified the transglutaminase as a 101 kDa monomeric protein with half-maximal cross-linking activity at 0.7mM Ca2+. The transglutaminase is also an ATPase and GTPase with apparent Km values of 15 mM ATP and 30 mM GTP.  Whereas Ca2+ is a positive effector for the cross-linking reaction, it is an inhibitor for the ATPase and GTPase activities. Conversely, purine nucleoside di- and triphosphates are negative effectors for the cross-linking activity. 

The Physarum transglutaminase appears analogous to human tissue transglutaminase which is found in all organs and believed to have roles in apoptosis, -adrenergic signal transduction, and possibly degenerative brain diseases such as Huntington's.  However, few specifics are known about its functions in man.  To better understand its role in the model organism Physarum we are characterizing cross-linking target proteins. LAV1-2, a plasmodium specific 40-kDa protein with four EF-hand calcium-binding motifs, is the preferred in situ cytoplasmic acyl donor.  In vitro cross-linking with purified transglutaminase and proteins fractionated by DEAE chromatography indicates that LAV1-2 is conjugated to a small set of acyl acceptor proteins. 

Figure 1: Ghost Plasmodia at Three Stages of the Cell Cycle

GhostPlasmodiaatThreeStagesoftheCellCycle

Antibodies raised against cross-linked ghost structures have been used to select target protein clones from a cDNA library.  These antibodies lightly label cytoplasm of plasmodia but strongly decorate nucleoplasm in a punctate pattern (Figure 2). On immunoblots the antibodies strongly bind seven nuclear and two cytoplasmic proteins. Among the interesting target proteins are Nuclear Ghost Protein 1 (NGP1, comprised of a central domain with two RNP1-type RNA recognition motifs, flanked by N-terminal lysine and glutamate-rich regions predicted to form helical and coiled coil structures.  At the C-terminus is a glycine-rich domain with GGRGGF repeats similar to many hnRNP proteins. The 61.1-kDa NGP2 contains a single RNP1 motif and possible CCHC zinc finger flanked at the N-terminus by 20 repeats with consensus ESGWGE(S/G)0-2GDK(E/EK/KE). NGP3 is a member of the UPF24 family of proteins predicted from ORFs in eubacteria, archaea, and eukaryotes but which are of unknown function.  

Figure 2: Nuclear Ghost Protein Immunofluorescence

NuclearGhostProteinImmunofluorescence

Publications

Mottahedeh, J., Leight, A., Sehgal, R., Vo, K., and R. Marsh, Physarum Transglutaminase and Its Substrates, 2000 Physarum Conference, Quebec, Canada 2000 - Publication
Mottahedeh, Jack, and Robert Marsh. Characterization of 101-kDa Transglutaminase from Physarum polycephalum and Identification of LAV1-2 as Substrate The Journal of Biological Chemistry 273, 29777-29895 1998 - Publication
Chang-Goo Huh, Joan Aldrich, Jack Mottahedeh, Heechung Kwon, Clayton Johnson, and Robert Marsh Cloning and Characterization of Physarum polycephalum Tectonins: Homologues of Limulus Lectin L-6. The Journal of Biological Chemistry 273, 6565-6574. 1998 - Publication
Mottahedeh, J., and R. Marsh, SDS-Insoluble Nuclear Ghosts of Physarum polycephalum: Fact or Artifact? 1996 European Physarum Meeting, , Norway 1996 - Publication
Aldrich, J., and R Marsh, Intracellular Location of the Tectonins: the 39 and 25 kDa Proteins in Physarum polycephalum. 1996 European Physarum Meeting, , Norway 1996 - Publication
Mottahedeh, J., and R. Marsh, SDS-Insoluble Nuclear Ghosts of Physarum polycephalum: Fact or Artifact, 1996 European Physarum Meeting, , Norway 1996 - Publication
Mottahedeh, J., Aldrich, J., and R. Marsh, Tectonins I and II are not nuclear matrix proteins but appear to be associated with cytoplasmic vesicles. 1994 European Physarum Meeting 1994 - Publication
Huh, C.-G., F. Yau, C. Johnson, and R. C. Tectonins I and II: Primary Structures and Genes. North American Physarum Conference, Cornell Univ., Ithaca, NY. Physarum Newsletter 21, No. 1, p. 28. 1989 - Publication
Stover, C.K., J. Kemper and R.C. Marsh, Molecular cloning and characterization of supQ/newD, a gene substitution system for the leuD gene of Salmonella typhimurium. The Journal of Bacteriology. 170, 3115-3124. 1988 - Publication
Johnson, C., C.-G. Huh, and R.C. Marsh, The matrix proteins tectonins I and II of Physarum nuclei and mitochondria share amino acid sequences with each other and with single strand DNA binding proteins. 8th European Physarum Workshop. Biologische Chemie Hoppe-Seyler 369, 1083. 1988 - Publication

Projects

Characterization of Histone-Depleted Nucleoli from Physarum polycephalum
1981–1981 1981, Kan, C.-C. and R. Marsh, Characterization of Histone-Depleted Nucleoli from Physarum polycephalum. Abstracts of the 81st Annual Meeting of the American Society for Microbiology, Abstract No. H27, p. 118.
Isolation and Characterization of the Escherichia coli origin of Replication
1978–1978 1978, Marsh, R. C., Isolation and Characterization of the Escherichia coli origin of Replication. 78th Annual Meeting of the American Society for Microbiology
Studies on the Relationship Between Replication and Expression of Phage T4 Late Genes
1971–1971 1971, Marsh, R.C, Studies on the Relationship Between Replication and Expression of Phage T4 Late Genes. 1971 European Phage Meeting Abstracts.
Molecular Structure of the Extrachromosomal Nucleolus of Physarum polycephalum
1983–1983 1983, Kan, C.-C. and R. Marsh, Molecular Structure of the Extrachromosomal Nucleolus of Physarum polycephalum. The Ninth North American Physarum Conference, Abstract No. XII.
Interaction of EF-G and EF-T with Ribosomes
1972–1972 1972, Parmeggiani, A., G. Chinali, R.C. Marsh, and G. Sander, Interaction of EF-G and EF-T with Ribosomes. Cold Spring Harbor Abstracts, Protein Synthesis Meeting, p. 49.

Additional Information

24 publications in related fields
COMMITTEE MEMBERSHIP AND OTHER UNIVERSITY SERVICE:
  • Ad hoc Committee on Periodical Reading Room for McDermott Library (1976)
  • Biology Graduate Education Committee (1979-1989)
  • Biology Undergraduate Education Committee (1989-), Chairman and Biology
  • Undergraduate Advisor (1994-)
  • Biology Faculty Search Committee (1979, 198l-1982, 1995, Chair 1996, Chair 1997)
  • Institutional Biosafety Committee (1979-98), Chairman (1989-1992), (Vice-Chair, 1997)
  • Biology MAT Education Committee (1980-1995), Chairman (1989-1995)
  • Committee on Research Involving Human Subjects (l980-l984)
  • Academic Senate (1986-1987, 2002-2005)
  • Radiation Safety Officer/Laser Safety Officer (1991-2002)
  • Radiation Safety Committee, Chair (1995-2002), Vice-Chair (2003-2005)
  • Health Professions Advisory Committee (1991-)
  • Advisory Committee for M.S. in Environmental Sciences (1998)
  • Committee on Educational Policy (1999-2001, 2003-2005) , Chair (2004-2005)
  • Compliance Subcommittee (2003-)
STUDENTS SUPERVISED - M.S. Awarded
  • William R. Parks (May) 1978
  • Jiing-Kuan Yee (December) 1979
  • Mariko Yamaguchi (May) 1980
  • Clifton L. Rule (May) 1981
  • Mary Jo Maher (Interim, May) 1981
  • Chen-Chen Jane Kan (May) 1981
  • Chi-Tang David Hsu (Interim, May) 1982
  • Wei-Marie Shen (non-thesis, May) 1983
  • Robert S. Kushner (Interim, May) 1985
  • Clayton H. Johnson (Interim, May) 1986
  • Seong Kyu Song (December) 1987
  • Chung-Goo Huh (Interim, May) 1988
  • Enrico A. Duru (May) 1989
  • Heechung Kwon (December) 1992
  • Jack Mottahedeh (Interim, May) 1994
  • Andre Frieden (December) 1994

Funding

Characterization of Tectonins and Tectonin- Related Proteins of Physarum polycephalum and Saccharomyces cerevisiae
$1,500 - National Institutes of Health Biomedical Research Support Grant RR07133-20 [1989–1990]
Characterization of Tectonins and Tectonin- Related Proteins of Physarum polycephalum and Saccharomyces cerevisiae
$1,500 - National Institutes of Health Biomedical Research Support Grant RR07133-19 [1988–1989]
"Characterization of Tectonins I and II: DNA-associated Proteins of Nucleoli, Nucleoplasm, Mitochondria, and Chloroplasts
$1,500 - National Institutes of Health Biomedical Research Support Grant RRO7l33-l8 [1987–1988]
In vivo Functionality of T4 Origins of Replication
$9,638 - UT-D Organized Research Grant l0655-072 and TPEG Scholarship funds [1985–1986]
Molecular Architecture of Nucleoli
$151,906 - National Science Foundation Grant PCM-83030l0 [1983–1987]