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Marzluff, William F.

June 14, 2011

My research interests are focused on the regulation of gene activity in animal cells, in particular regulation of gene expression during the cell cycle by postranscriptional mechanisms. One system we study is the regulation of histone mRNA, both during the mammalian cell cycle and during early development in frogs and sea urchins. Histone mRNAs are the only mRNAs that do not have polyA tails, ending instead in a conserved stem-loop structure. Histone mRNAs are present only in S-phase cells and most of the regulation is mediated by the 3′ end of histone mRNA. Histone genes lack introns, and histone mRNA is formed by a single processing reaction, cleavage to form the 3′ end of the histone mRNA. The mRNA is then immediately transported to the cytoplasm. Both the cleavage reaction to form histone mRNA and the half-life of the histone mRNA are regulated during the cell cycle. We have cloned the cDNA for the stem-loop binding protein (SLBP) that binds the 3′ end of histone mRNA and participates in all aspects of histone mRNA metabolism. SLBP is a critical factor involved in regulating histone mRNA levels. Our current interests are in understanding how SLBP carries out its multiple functions; RNA binding, 3′ processing, transport, stimulator of translation and regulating histone mRNA half-life. In addition, we are studying how SLBP itself is regulated and how this regulation connects the other cell cycle regulators with the regulation of histone mRNA.

Sea Urchin

In embryos which undergo a very rapid series of cell divisions after fertilization, there is an exponentially increasing demand for histones to assemble the newly replicated DNA into chromatin. During this time the histone mRNAs are not cell-cycle regulated but are stable for multiple cell cycles. We have cloned embryo specific SLBPs from these stages and are determining how they function to regulate histone mRNA metabolism in frog and sea urchin embryos. We are also studying the role of the G1 cyclins, cyclin D and cyclin E in the regulation of these early cell cycles that lack gap phases.

Matera, Gregory

June 14, 2011

At a Glance

  • Epigenetic gene regulation
  • Animal models of Spinal Muscular Atrophy
  • RNA processing and RNP assembly

The research in our laboratory interrogates fundamental molecular, cellular and developmental biological processes. In particular, we are interested in the roles played by small ribonucleoproteins (RNPs) in a genetic disease, called Spinal Muscular Atrophy.  We are also focused on the functions of histone post-translational modifications (PTMs) in the regulation of eukaryotic gene expression, important for understanding disease mechanisms in many different types of cancer.

 

Epigenetics (Regulation of Gene Expression)

  • Role of histone PTMs in transcription and RNA processing
  • Understanding contribution of histone PTMs to cellular memory
  • Developing new tools to study multi-copy gene families

 

SMA (Spinal Muscular Atrophy)

  • Function of the SMN complex in snRNP biogenesis
  • Use of animal models to understand disease etiology
  • Identification of novel functions of SMN complex

Reed, Jason W.

June 2, 2011

Most plant development occurs post-embryonically, and is tied closely to environmental signals.  We use techniques of genetics, molecular biology, microscopy, physiology, and biochemistry to study how environmental and endogenous signals regulate plant development.  For these studies we use the model plant Arabidopsis thaliana, which has numerous technical advantages that facilitate experimental progress.  We hope in the long run to reconstruct how endogenous developmental programs and exogenous signals cooperate to determine plant form and the partitioning of growth among different organs.

The hormone auxin regulates multiple developmental processes in plants, including embryo and meristem patterning, organ growth, and flower maturation.   Auxin induces gene expression through a family of transcription factors called ARFs (Auxin Response Factors), whose activity is regulated by Aux/IAA proteins.  Auxin switches ARF proteins between gene repressing and gene activating states by promoting Aux/IAA protein turnover.  We are currently studying how this regulatory system controls seedling growth, ovule development, and flower opening and fertilization.

Among genes regulated by auxin are some (including those encoding Aux/IAA proteins) that feed back negatively on auxin response; and others encoding proteins that affect intercellular auxin transport.  We are interested in how feedback controls affect the dynamics of auxin response, and in how regulated intercellular auxin movement coordinates growth and differentiation of different cells during development.

 

Duronio, Robert J.

May 25, 2011
Drosophila embryonic cells (purple) undergoing mitosis (green).

At a Glance

  • Epigenetic control of genome structure and function
  • Cell cycle-regulated gene expression
  • Developmental genetics

Synopsis

Our research focuses on understanding the molecular mechanisms that regulate DNA replication and cell proliferation during animal development.  An orderly process of events called the cell cycle, which in its most familiar form consists of four phases (G1-S-G2-M) controls cell proliferation.  The genome is replicated during the “S” or synthesis phase and duplicated chromosomes are segregated to daughter cells during the “M” or mitotic phase when cell division occurs.  G1 and G2 are “gap” phases during cells regulate the entry into S phase and M phase, respectively.  We study gene expression events that control how cells make the decision to enter S phase and proliferate, or to exit the cell cycle and differentiate. Without such control there would be no coordination between cell proliferation and the development and function of the many different types of tissues that make up an organism.  In addition, the breakdown of cell cycle regulation is one of the events that contribute to the generation of cancer.

The E2F transcription factor is active (green) during S phase (red) in the asynchronous endocycles of the Drosophila salivary gland.

We study this problem using the fruit fly Drosophila melanogaster, in part because the genes controlling cell proliferation in fruit flies have been highly conserved during evolution and function the same way as in other animals, including humans.  This allows us to exploit certain advantages that Drosophila has as a research tool, including the ease with which genetics (making and analyzing mutants) and cell biology (using microscopy to

In situ hybridization for histone gene expression in a Drosophila embryo.

observe cell proliferation) can be applied to the study of gene function in the context of a whole animal.  Some of the cell cycle regulatory pathways that we study become defective in virtually every human cancer.  Thus, one hope is that understanding how these pathways function in normal Drosophila development will give us clues to how they might malfunction in the deregulated growth typical of cancer.

Conlon, Frank

May 20, 2011

Human congenital heart disease, the most common form of heart disease in childhood, occurs in about 1% of live births and up to 10% of stillbirths. Presently, the most effective therapy for cardiac diseases is heart transplantation. However, due to the shortage of organs, cost and inaccessibility of treatment for most affected individuals this remains a limited therapeutic option. Alternative treatment is the administration of drugs that improve myocardial contractility, though this treatment is only effective as a short term therapy, with the 5-year survival rate using current agents being less than 60%. An alternative therapeutic option is to treat patients with cardiac progenitor cell populations that could infiltrate and repair damaged heart tissue. Thus, the ability to isolate and propagate cell populations that can differentiate into cardiomyocytes in vivo offers the opportunity to treat a wide range of cardiac diseases. To this end, our lab is interested in understanding the relationship between cardiac progenitor proliferation and the onset of cardiac differentiation focusing on the endogenous roles of the transcription factors TBX5 and CST and the protein phosphatase SHP-2.

Crews, Stephen

April 21, 2011

At a Glance

  • Formation and differentiation of Drosophila neurons and glia
  • Regulation of nervous system transcription

Synopsis
Our laboratory is concerned with the molecular mechanisms that govern the development of the Drosophila central nervous system, including: (1) how nervous system precursor cells are generated, and (2) how neurons and glia acquire their differentiated properties. The primary focus of the lab is using genomic technologies to study the regulation of transcription during neuronal and glial development.

Bautch, Victoria L.

April 13, 2011

We study the growth and interactions of cells in their natural environment – the animal – and how these interactions are modified in disease. We focus on the mechanisms that control the process of blood vessel formation, which is crucial to successful development and required in cancer and other diseases. There are currently several areas of investigation that utilize genetically altered mice and cells derived from those mice.

  • We developed a cell culture model of developmental blood vessel formation to study cross-talk between cellular processes such as cell division and sprouting migration to expand vessel networks. Mouse embryonic stem cells are induced to differentiate in dishes to form structures that contain some embryonic tissues, including primitive blood vessels. We have incorporated GFP reporter genes to visualize the dynamic processes of blood vessel formation via time-lapse imaging, and we have used both genetic manipulation and inhibitors to dissect the role of an important signaling pathway (VEGF) in these processes.
  • We study the role of a novel gene that activates cellular homologs of oncogenes (ie Ras) in the proper function of blood vessels. We showed that while this gene is not required for development, it is required for the response of vessels to phorbol esters, which are tumor promoters. This suggests that the signaling pathway using this gene is important in diseases such as diabetes and cancer, and we are testing these models.
  • We investigate how blood vessels know where to go as they form, by using chimeric embryos that consist of a host bird embryo with inserted mouse tissue. This technique allows us to determine the migration patterns of blood vessels over time during fetal development, and we can genetically manipulate the mouse tissue. We can also introduce exogenous DNA into the host via electroporation of the embryos. This analysis has thus far uncovered a crucial role for VEGF in the patterning of vessels around the neural tube, which will form the brain and spinal cord.

Ahmed, Shawn

April 12, 2011

Our research group has interests in telomere biology and germline immortality. We are studying these problems using the nematode Caenorabditis elegans, a multicellular eukaryote with excellent genetics, which can be combined with biochemical and cell biology approaches.

How does the germline achieve immortality?

The germline is an immortal cell lineage that is passed from one generation to the next, indefinitely. In order to determine how germ cells achieve immortality, we have identified C. elegans mutants with Mortal Germlines: mutants that reproduce normally for several generations, but eventually became sterile (Figure 1). Gametes of these mutants transmit forms of stress that can accumulate over the generations to levels that cause strong effects on germ cell development (sterility or reproductive arrest). Stress transmitted by mortal germline mutants could also affect somatic development. One pathway that promotes germ cell immortality is the Piwi/piRNA genome silencing pathway. Deficiency for this pathway allows for growth for many generations but then results in an adult reproductive arrest phenotype that resembles a response to acute starvation. Furthermore, deficiency for the daf-2 insulin/IGF1 receptor, which results in stress resistance and adult longevity, can suppress the sterility of Piwi mutants. This creates an interesting link between germ cell immortality and somatic longevity whose basis we seek to understand.

Figure 1. mortal germline mutants eventually become sterile after several generations.

Given that the soma is only needed for a single generation, evolutionary theory posits that somatic cells may be deficient for pathways that ensure germ cell immortality. Substantial evidence from vertebrates indicates that this is likely to be the case. We utilize genetic, cell biology and biochemical approaches to study pathways that enable germ cells to achieve proliferative immortality in C. elegans. This work could shed light on epigenetic forms of macromolecular stress that can be transmitted by gametes and could be generally relevant to heredity and possibly to human disease.

Analysis of Telomere Biology

A second pathway that promotes germ cell immortality is telomerase, which is a ribonucleoprotein that adds repeats to chromosome termini. The ends of linear chromosomes, telomeres, are usually composed of simple repetitive sequences whose length is maintained by telomerase. Humans deficient for telomerase suffer from the lethal hereditary disorders Aplastic Anemia, Dyskeritosis Congenita and Pulmonary Fibrosis. C. elegans is the most highly evolved multicellular eukaryote in which an unbiased genetic approach can be used to study the problem of telomere replication. A fraction of the mortal germline mutants described above are completely defective for telomerase activity in vivo (Figure 2). Studies of these genes may help to elucidate the mechanism by which telomerase functions at chromosome termini and may provide significant insight into telomerase-related diseases. We also study proteins that bind to telomeres, some of which form nuclear foci that are dynamic during development.

Telomerase is deficient in most human somatic cells, and telomere erosion in this context may contribute to genome instability that fuels the development of many forms of cancer. C. elegans has holocentric chromosomes, so fused chromosomes that occur when telomerase is deficient can be genetically isolated and mapped. These stable end-to-end chromosome fusions allow us to address the mechanism by which dysfunctional telomeres are repaired with unparalleled clarity, and may provide insight into how large-scale DNA rearrangements occur during tumorigenesis.

Figure 2. Teomere replication defects such as (a) telomere shortening and (b) end-to-end chromosome fusions are observed in a subset of mortal germline mutants, suggesting that teomerase does not function properly in these mutants.