# GLYCOCONJUGATES IN DEVELOPMENT AND IMMUNITY

> **NIH NIH R01** · UNIVERSITY OF ALABAMA AT BIRMINGHAM · 1994 · $234,886

## Abstract

The long-term goal of this project remains the elucidation of the
biological functions of glycosyl moieties on glycoconjugates important
to development and immunity.  During studies on lymphocytes, a new type
of protein glycosylation (O-GlcNAc) was discovered.  O-GlcNAc occurs in
all eukaryotes from yeast to man on a myriad of nuclear and cytoplasmic
proteins, including RNA polymerase II and its transcription factors,
numerous chromatin-, nuclear pore-, cytoskeletal-, viral-, nuclear
oncogene-, and heat shock-proteins.  O-GlcNAc appears to be ubiquitous,
abundant, and as dynamic as phosphorylation.  Elucidation of O-GlcNAc's
functions will be the focus of this project's next phase.

Our plan is to:  1) Complete the characterization, clone the genes, and
understand the regulation, of the enzymes that attach or remove O-GlcNAc.
Complexity of gene families, regulation of the enzymes at the genomic,
mRNA, and protein levels will be determined.  Recombinant enzymes will
be used to develop inhibitors, and to study the enzymes' structures.  2)
Several approaches will evaluate the general significance of O-GlcNAc:
A. CHO-mutant - Cytoplasmic constructs of Gal transferase cDNA will be
transfected into gal-deficient CHO cells (LDL-d), and the deleterious
effects of exogenous galactose on viability, growth, nuclear transport,
transcription, and other properties will be evaluated.  Preliminary data
suggests that similar cytoplasmic constructs are lethal in wild-type
cells.  B. Yeast - O-GlcNAc transferase cDNA from yeast will be cloned.
Classical gene knock-out experiments will be performed.  Effects of
glucosamine deprivation on O-GlcNAc will be studied in a yeast Ts
auxotrophic for GlcN.  C. Inhibitors - Inhibitors of O-GlcNAc
transferase, such as polyoxins, and inhibitors of O-GlcNAcase, such as
NAc-deoxynorjirimycin, will be studied.  D. Anti-Sense - Overexpression
of anti-sense cDNA for O-GlcNAc transferase and O-GlcNAcase will also be
attempted to specifically effect O-GlcNAc metabolism.  3) Determine the
relationship between O-GlcNAc and Ser/Thr phosphorylation:  Initially,
we will co-examine synthetic peptide substrates with pure O-GlcNAc
transferase and purified kinases.  Co-localization of sites on endogenous
proteins and effects of phosphatase/kinase inhibitors on O-GlcNAc and
vice versa will be determined.  4) Continue to Identify O-GlcNAc
Attachment Sites for Site-Directed Mutagenesis:  Initially, we will focus
on RNA polymerase II, serum response and HNF-1 transcription factors, and
Neurofilaments.  5) Dynamics of O-GlcNAc will be further explored in
cellular activation, cell cycle, and developmental systems.  Overall,
these experiments will elucidate the functions of O-GlcNAc, a ubiquitous,
abundant, and dynamic post-translational modification, which may have
substantial impact on nearly all aspects of eukaryotic biology, including
development and intracellular processes.

## Key facts

- **NIH application ID:** 2197016
- **Project number:** 5R01HD013563-16
- **Recipient organization:** UNIVERSITY OF ALABAMA AT BIRMINGHAM
- **SAM.gov UEI:** YND4PLMC9AN7
- **Principal Investigator:** GERALD Warren HART
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1994
- **Award amount:** $234,886
- **Award type:** 5
- **Project period:** 1979-12-01 → 1997-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/2197016

## Citation

> US National Institutes of Health, RePORTER application 2197016, GLYCOCONJUGATES IN DEVELOPMENT AND IMMUNITY (5R01HD013563-16). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/2197016. Licensed CC0.

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