Timothy C. Umland, Ph.D.
Research Scientist, Hauptman-Woodward Inst.
Assistant Professor of Structural Biology, SUNY-Buffalo

EDUCATION
B.S. in Chemistry, SUNY-Buffalo, 1988
Ph.D. in Crystallography, University of Pittsburgh, 1994

MAILING ADDRESS:
Hauptman-Woodward
Medical Research Institute
700 Ellicott Street
Buffalo, NY 14203-1102

CONTACT INFORMATION:
Tel. 716-898-8632
Fax. 716-898-8660
Email. umland@hwi.buffalo.edu

Research Interests

Structural determinants of DNA-binding  specificity by HOX and HOX-cofactors. Coronavirus proteins and host range expansion. 3β-hydroxysteriod dehydrogenase.

Structural determinants of DNA-binding specificity by HOX and HOX-cofactors

The HOX transcription factor proteins are master regulators intimately involved in controlling cellular proliferation and differentiation in all multicellular animals exhibiting bilateral symmetry. During the development of a human embryo to a fetus, the human HOX proteins control development along the anterior-posterior axis. That is, they control what part of the body becomes the head, becomes the arms, becomes the legs, becomes the toes, and so on. The HOX family also controls the proper development of the internal organs (e.g., central nervous system, heart, kidney, etc.). In short, the HOX family is instrumental to a human looking like a human, a mouse like a mouse, and a jellyfish like a jellyfish. Throughout life the HOX family is involved in controlling the formation of new blood cells (hematopoiesis) through the proliferation and differentiation of hematopoietic stem cells. HOX mutations have been associated with cancer, including several types of leukemia.

HOX proteins act as master regulators through their ability to specifically turn on or off a wide variety of genes. Surprisingly, very few of these target genes have actually been identified. Each of the thirty-nine members of the human HOX family exhibits different biological activity, and a precise balance must be maintained for proper development to occur. However, it is unknown how each of these related proteins control different sets of genes. Furthermore, the identity of very few human genes that are regulated by the HOX family is known, despite the established developmental importance of these proteins in both humans and model organisms. In part, this is due to the inability to accurately predict regulated genes based upon regulatory DNA (promoter) sequence, as core DNA sequences that bind to HOX proteins in vitro often fail to act as regulatory sequences in vivo. Our collaborator (Kenneth Gross, Roswell Park Cancer Institute) has identified the human renin gene as being regulated by HOX proteins. We are using this genuine HOX regulatory sequence to investigate the interactions between HOX proteins, DNA, and other protein partners (cofactors) that form complexes that can either enhance or suppress gene expression. My lab uses X-ray crystallography to examine these interactions at high resolution. We also use other biophysical methods to characterize these protein•DNA interactions. HOX proteins also bind to other proteins (cofactors) that have the ability to alter DNA specificity and gene regulatory activity. In collaboration with both the Gross lab and Steven Pruitt's lab (Roswell Park Cancer Institute), we are identifying new HOX cofactors, and targeting them for structure-function studies.

SARS-CoV and Coronavirus Proteins

The Severe Acute Respiratory Syndrome (SARS) outbreak of 2002-2003 followed by a smaller outbreak in 2004 killed over 800 people and caused massive financial losses worldwide. It prompted a renewed interest in coronavirus (CoV) research. Prior to the SARS outbreak, only two CoVs (HCoV-229E and HCoV-OC43) were known to infect humans. These two CoVs have been estimated to cause 30% of common colds. CoVs are widespread in both domestic and wild animals, with several being major problems in farm animals (pigs, cattle, chickens). Mouse hepatitis virus (MHV) is widespread among mice in research facilities, and MHV infection can significantly alter research studies.  Following the emergence of SARS, two additional CoVs were identified (NL63 and HKU1). NL63 is widespread in children and usually causes a mild respiratory disease. However, occasionally it causes severe respiratory infection in young children. NL63 has likely been present in humans for some time, and only recently recognized due to improved technologies and renewed interest in CoVs.

In collaboration with Dr. Wayne Schultz (HWI), my lab began studying SARS-CoV shortly after SARS-CoV was identified in Toronto (90 miles away from HWI). We have pursued several research directions: (a) development of a passive immunotherapy, (b) studies of the viral proteins involved in viral replication and pathogenesis, and (c) virus•host protein•protein interactions influencing virulence and host species selection.  The passive immunotherapy project is in collaboration with ZeptoMetrix (Buffalo, NY) and Virionyx (Aukland, NZ). Antibodies raised in goats against inactivated virus or viral proteins can be used to treat newly infected people in a manner analogous to snake anti-venom. The CoV has a positive-strand RNA genome, which is the largest for any RNA virus. The CoV family has proteins which are not found outside of its family, and this may be because proteins evolved within the CoV family to maintain its large and inherently error prone RNA genome. Structural and biochemical characterization of these novel viral replicase proteins will reveal how RNA viruses can stable expand their genomes to acquire new and more complex functions, and may provide new targets for therapeutic drugs. My lab has recently begun to explore interactions between SARS CoV proteins and human proteins. Virus•host interactions are required for viral replication, virion formation, immunosupression, and adaptation to a new host. Despite the importance of virus•host interactions, relative few are well characterized. In collaboration with Dr. Steven Pruitt (RPCI), we are undertaking a comprehensive screen of all SARS-CoV•human protein•protein interactions. We will compare these interactions to those between civet-SARS CoV and human proteins, and those between bat-SARS CoV and human proteins in order to identify the interaction differences attributable to host range expansion from the animal reservoir to humans.

3β-Hydroxysteriod Dehydrogenase/Isomerase

Type 1 and type  2 isoforms of 3β-hydroxysteriod dehydrogenase (3β-HSD1 and 3β-HSD2) are also known as 3β-hydroxy-D⁵-steroid dehydrogenase (EC 1.1.1.145)/3-oxosteroid D⁵-D⁴-isomerase (EC 5.3.3.1) types 1 and 2. This more descriptive name indicates the dual enzymatic reactions that the protein catalyzes. The two isoforms are encoded by distinct genes in humans, and are expressed in a tissue specific pattern (3β-HSD1: placenta, skin, mammary gland, prostate, endometrium; 3β-HSD2: gonads, adrenals). Both isoforms of 3β-HSD perform early steps in the synthesis of important steroidal hormones. In human placenta, 3β-HSD1 catalyzes the conversion of  3-hydroxy-5-ene steroids (dehydroepiandrosterone, pregnenolone) to  3-oxo-4-ene steroids (androstenedione, progesterone) on a single, dimeric protein containing both enzyme activities. Androstenedione is then converted by placental aromatase and 17-hydroxysteroid dehydrogenase  (17-HSD) to estradiol, which participates in the cascade of events that  initiates labor in humans. In addition to placenta and  other human peripheral tissues, the 3β-HSD1 enzyme is selectively expressed in  breast tumors, prostate  tumors, and  choriocarcinomas, where it  catalyzes the first step in the conversion of circulating  dehydroepiandrosterone to estradiol or testosterone to promote tumor growth.  In human adrenals, type 2 3-HSD/isomerase is a key enzyme required for  the production of cortisol and aldosterone. The two isoforms have highly similar sequences (93% identical) and yet exhibit differing enzyme kinetics for both the dehydrogenase and isomerase reactions. Determination of the structure/function relationships of the type 1 enzyme may lead to the development of specific inhibitors of 3β-HSD1 that can  help control the timing of labor and slow the growth of hormone-sensitive  tumors without compromising the essential functions of the 3β-HSD2 adrenal enzyme, and thus significantly lessening the side effects. We are conducting these structure/function studies in collaboration with Dr. James Thomas (Mercer University School of Medicine) and Dr. William Duax (HWI).

 The two-step reaction of 3β-HSD using dehydroepiandrosterone  (DHEA) as substrate is shown in below. This reaction scheme shows the reduction of NAD+ to NADH by the rate-limiting dehydrogenase activity and the requirement of this NADH for the activation of isomerase on the same enzyme protein

Selected Publications

• Brucz K, Miknis ZJ, Schultz LW, Umland TC. (2007). Expression, purification and characterization of recombinant severe acute respiratory syndrome coronavirus non-structural protein 1. Protein Expr Purif., 52, 249-57.
• Mao Q, Duax WL, Umland TC. (2007). Crystallization and X-ray diffraction analysis of the beta-ketoacyl-acyl carrier protein reductase FabG from Aquifex aeolicus VF5. Acta Crystallograph Sect F Struct Biol Cryst Commun., 63, 106-9.
• Pletnev VZ, Thomas JL, Rhaney FL, Holt LS, Scaccia LA, Umland TC and Duax WL. Rational Proteomics V: Structure-based mutagenesis has revealed key residues responsible for substrate recognition and catalysis by the dehydrogenase and isomerase activities in human 3beta-hydroxysteroid dehydrogenase/isomerase type 1.
  J. Steroid.Biochem. Mol. Biol., 101, 50-60.
• Brody L, Cookson B, Fletcher, DA, Gerrard S, Lorenz M, Loudermilk A, Okie S, Orbach M, Singh M, Subramaniam S and Umland T. Working Group Summary: Are there shared pathways of attack that might provide new avenues of prevention?, in The Genomic Revolution: Implications for Treatment and Control of Infectious Disease. National Academies Keck Futures Initiative, National Academies Press, Washington, D.C. (2006).
• Thomas JL, Boswell EL, Scaccia LA, Pletnev V and Umland TC. Identification of key amino acids responsible for the substantially higher affinities of human type 1 3β-hydroxysteroid dehydrogenase/isomerase (3β-HSD1) for substrates, coenzymes, and inhibitors relative to human 3β-HSD2. J. Biol. Chem., 280, 21321-21328 (2005).
• Thomas JL, Umland TC, Scaccia LA, Boswell EL and Kacsoh B. The higher affinity of human type 1 3β-hydroxysteroid dehydrogenase (3β-HSD1) for substrate and inhibitor steroids relative to human 3β-HSD2 is validated in MCF-7 tumor cells and related to subunit interactions. Endocr. Res., 30, 935-941 (2004).
• Umland TC, Wolff EC, Park MH and Davies DR. A new crystal structure of deoxyhypusine synthase reveals the configuration of the active enzyme and of an enzyme•NAD•inhibitor ternary complex. J. Biol. Chem., 279, 28697-28705 (2004).
• Umland TC, Taylor KL, Rhee S, Wickner RB and Davies DR. The Crystal Structure of the Nitrogen Regulation Fragment of the Yeast Prion Protein Ure2p. Proc. Natl. Acad. Sci.,98, 1459 -1464 (2001).
• Umland TC, Wei S-Q, Craigie R and Davies DR. Structural Basis of DNA Bridging by Barrier-to-Autointegration Factor. Biochemistry, 39, 9130-9138, (2000).
• Umland TC, Wingert LM, Swaminathan S, Furey WF, Schmidt JJ, and Sax M. The Structure of the Receptor Binding Fragment HC of Tetanus Neurotoxin. Nature Struct. Biol., 4, 788-792, (1997).