LINDA M. HENDERSHOT
ASSISTANT PROFESSOR

EDUCATION:
B.S. 1975, Eastern Kentucky University, Richmond
Ph.D. 1983, University of Alabama at Birmingham
Postdoctoral 1983-1985, University of Alabama at Birmingham

RESEARCH INTERESTS: In the ER, nascent polypeptides must fold and assemble in an oxidizing milieu containing millimolar concentrations of protein and of calcium, each of which inhibits protein folding in vitro. Cellular proteins known as molecular chaperones have been shown to facilitate protein folding by genetic means and in vitro assays, but in many cases their in vivo interactions remain unclear. Based primarily on their association with folding or assembly intermediates, several ER protein have been proposed to function as molecular chaperones. The best characterized is BiP/GRP78, an hsp70 homologue that binds peptides containing hydrophobic residues in vitro and unfolded or unassembled proteins in vivo. All the Er chaperones are transcriptionally upregulated in response to the accumulation of unfolded proteins in the ER. This coordinately regulation involves monitoring the level of BiP-associated proteins and transducing a signal across the ER membrane to increase or decrease the transcription of the ER chaperones, and have initiated experiments to understand the signal transduction pathway from the ER to the nucleus that controls the transcriptional level of these genes.
The ER "molecular chaperones" include BiP (the ER hsp70 cognate), grp94, Erp72, and calnexin. Currently, it is unclear if they act only to prevent misfolding and incorrect assembly, or if they also play a more active role in these processes. Moreover, the relationships and interactions between the various ER chaperones have not been established. We have focused our attention on BiP because it may represent the initial chaperone encountered by a nascent polypeptide upon entering the ER. We hypothesize that BiP is an essential component of the folding and assembly apparatus and that expression of inactive BiP mutants will have profound effects on the interaction of target proteins with other chaperones and ultimately in the maturation and secretion. The aims of our current studies are to define the nature of BiP's interaction and nascent proteins, to establish the temporal relationship of this interaction with that of other ER chaperones, to determine whether interactions exist between the various ER chaperones, and to establish the requirements for a newly synthesized protein to successfully navigate the ER quality control machinery.
In order to identify ER signals that initiate the signal transduction pathway that regulates the transcription of all ER chaperones, we investigated the role of BiP in the pathway. Chinese hamster cells were transfected with a hamster BiP cDNA clone and lines that constitutively express high levels of BiP from an adenovirus promoter were isolated. Forced expression of BiP from a heterologous promoter )1 down-regulates the endogenous expression of all of the ER chaperones, and 2) blocks their induction in response to malfolded proteins in the ER. This suggests that levels of BiP are monitored and used to regulate (both positive and negative) the transcription of the ER chaperones. Very recently, we have found that the growth arresting, C/EBP homologous transcription factor, CHOP, is in the BiP-induction pathway. Our data demonstrate that the cellular response to a variety of physiological stresses is two fold: 1) cell growth is inhibited until the condition subsides, and 2) ER chaperones are transcriptionally upregulated to protect the cell. Our preliminary data suggest that both responses occur through a single pathway that is initiated in the endoplasmic reticulum.

CURRENT RESEARCH SUPPORT:
NIH R01 "Role of Molecular Chaperones in Ig Biosynthesis" April 1, 1996 - March 31, 2001; P.I. Linda M. Hendershot, 50% effort; direct costs for 1 year: $178,908, pending.
ACS "Control of Cell Growth: Signal Transcuction from the ER to the Nucleus" July 1, 1996 - June 30, 1999; P.I. Linda Hendershot, 40% effort; direct costs for 1 year: $88,104, pending.

PUBLICATIONS: Only selected publications appearing since 1988 are listed.
Pollok, B.A., Anker, R., Eldridge, P., Hendershot, L., and Levitt, D. (1988) Molecular basis of the cell surface expression of immunoglobulin m chain without light chain in human B lymphocytes. Proc. Natl. Acad. Sci. USA, 84, 9199-9203.
Hendershot, L.M. and Kearney, J.F. (1988) A role for human heavy chain binding protein in the developmental regulation of immunoglobulin transport. Mol. Immunol., 25, 585-595.
Hendershot, L.M., Ting, J., and Lee, A.S. (1988) Identity of the immunoglobulin heavy chain binding protein with the 78,000 dalton glucose regulated protein and the role of post-translational modifications in its binding function. Mol. Cell Biol., 8, 4250-4256.
Kerr, W.G., Cooper, M.D., Feng, L., Burrows, P.D., and Hendershot, L.M. (1989) Mu heavy chains can associate with a pseudo-light chain complex (yL) in human pre-B cell lines. International Immunology, 1, 355-361.
Ma, J., Kearney, J.F., and Hendershot, L.M. (1990) Association of transport-defective light chains with immunoglobulin heavy chain binding protein. Mol. Immunol., 27, 623-630.
Hendershot, L.M. (1990) Immunoglobulin heavy chain and binding protein complexes are dissociated in vivo by light chain addition. J. Cell Biol., 111, 829-837.
Freiden, P.J., Gaut, J.R., and Hendershot, L.M. (1992) Interconversion of three differentially modified and assembled forms of BiP. EMBO J., 11, 63-70.
Gaut, J.R. and Hendershot, L.M. (1993) Mutations within the nucleotide binding site of immunoglobulin-binding protein inhibit ATPase activity and interfere with release of immunoglobulin heavy chain. J. Biol. Chem., 268, 7248-7255.
Gaut, J.R. and Hendershot, L.M. (1993) The immunoglobulin-binding protein in vitro autophos-phorylation site maps to a threonine within the ATP-binding cleft but is not a detectable site of in vivo phosphorylation. J. Biol. Chem., 268, 12691-12698.
Gaut, J.R. and Hendershot, L.M. (1993) The modification and assembly of proteins in the ER. Curr. Opin. Cell Biol., 5, 589-595.
Hendershot, L.M., Valentine, V.A., Lee, A.S., Morris, S.E., and Shapiro, D.N. (1994) Localization of the gen encoding human BiP/GRP78, the endoplasmic reticulum cognate of the HSP70 family, to chromosome 9q34. Genomics, 20, 281-284.
Hendershot, L.M., Wei, J-Y., Gaut, J.R., Lawson, B., Freiden, P.J., and Murti, K.G. (1995) In vivo expression of BiP ATPase mutants results in disruption of the endoplasmic reticulum.
Mol. Biol. Cell., 6, 283-296.
Fitts, M.G., Metzger, D.W., Hendershot, L.M., and Mage, R.G. (1995) The rabbit B cell antigen receptor is noncovalently associated with unique heteromeric protein complexes: possible insights into the membrane IgM/IgD coexpression paradox. Mol. Immunol., 32, 753-759.
Wei, J-Y. and Hendershot, L.M. Characterization of the nucleotide binding properties and ATPase activity of recombinant hamster BiP purified from bacteria. J. Biol. Chem., in press.
Wei, J-Y., Gaut, J.R., and Hendershot, L.M. In vitro dissociation of BiP:peptide complexes requires a conformational change in BiP after ATP binding but does not require ATP hydrolysis.
J. Biol. Chem., in press.
Hendershot, L.M., Wei, J-Y., Gaut, J.R., Melnick, J., Aviel, S., and Argon, Y. Inhibition of immunoglobulin folding and secretion by dominant negative BiP ATPase mutants. Submitted.