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Nikolaus S. Trede

Assistant Professor of Oncological Sciences and of Pediatrics

B.S. Albert Ludwigs University Medical School, Germany

M.D. Albert Ludwigs University Medical School, Germany

Ph.D. Université Paris VII

Research

References

nikolaus.trede@hci.utah.edu

Nikolaus Trede's Lab Page

Research

The study of mice and patients with immune deficiencies has greatly enhanced our understanding of the molecular processes involved in developmental aspects of the immune system. However, mammalian immunology is handicapped by the fact that the earliest steps in the ontogeny of the immune system occur in-utero, and are therefore difficult to study in an in-vivo system. For example, mutants of GATA-3, a transcription factor of pivotal importance for T cell development, die in-utero due to an accompanying brain hemorrhage. Prominent among these early steps are thymus organogenesis and T cell development, crucial interdependent processes that establish a functional vertebrate immune system. Current understanding of vertebrate thymus development during embryogenesis remains incomplete and the genetic basis for many disorders of the immune system, including immunodeficiencies and autoimmune disorders remain to be elucidated, creating the need for novel approaches.

The zebrafish has emerged as a powerful vertebrate model system for genetic and pharmacologic screens. Given its small size and prolific fecundity, the zebrafish is ideally suited for vertebrate forward genetic screens. These types of screens are phenotype-driven, and mutants with phenotypes of interest are positionally cloned. This process has been greatly aided by the completion of the zebrafish genome sequencing project by the Sanger Center. Transgenic fluorescent lines of zebrafish are versatile tools that facilitate read-out of screens for mutants and biologic modifiers. in-vivo imaging in adult zebrafish is feasible, allowing detection of internal organs such as the pancreas. Furthermore, the immune system of zebrafish closely resembles that of mammals (ref. 2), and visualization of T cells can be accomplished using the lck-promoter::eGFP transgenic line we have generated in the past (ref.3). Tracking green fluorescent lymphocytes has also enabled us to establish a zebrafish model of T cell leukemia (ref. 1).

The focus of our lab is centered on modeling human diseases of the immune system in zebrafish. The key questions we address are: What is the genetic basis of immune ontogeny (when this process goes awry it can result in immunodeficiency), and what are the genes responsible for preventing uncontrolled activation (autoimmunity) or growth (leukemia) of lymphocytes? To answer these questions, we employ a variety of tools, including genetic screens using transgenic lines of zebrafish.

We have initiated screens in zebrafish looking for defects in adaptive. For adaptive immunity, the lineage of choice is the T cell, because the organ in which it develops, the thymus, is a superficial organ that can be readily uncovered by wholemount in-situ hybridization (WISH) or fluorescent microscopy. In addition, by looking for T cell specific gene expression in the zebrafish thymus, these screens pick up not only mutants of T cell, but also of thymus development. Mutants that have come out of our initial WISH screen using the Rag-l probe fall into two categories: Class I mutants have no obvious defects in other organs, Class II mutants have abnormalities of the pharyngeal arches. We have positionally cloned the first two mutants (ref.4), and are in the process of cloning the remaining mutants.

We have begun a new screen using the lck-promoter::eGFP transgenic line, where we will read out absence of T cells on day 6 of development by in-vivo fluorescent microscopy. Similarly, older mutated individuals will be screened for organ infiltration by fluorescent T cells (autoimmunity) or uncontrolled proliferation of T cells in the thymus (leukemia). In this screen we have already identified mutants with immunodeficiency and dominant, heritable mutations that lead to leukemic phenotypes. These mutants undergo rigorous secondary analysis and interesting mutants will be positionally cloned.

In collaboration with the HCI labs of David Jones and Brad Cairns we have also begun to analyze genome-wide methylation signatures in immature, mature and leukemic lymphocytes. This novel approach is designed to uncover epigenetic processes involved in normal and malignant development of immune cells.

References