Susan Strome - US grants

The establishment of asymmetry during early embryogenesis and the distribution of different developmental information to different embryonic cells are crucial processes in the development of every multicellular organism. Yet the nature of embryonic informational molecules and the mechanisms governing their differential distribution are not understood. This proposal delineates a combined immunologic, genetic, and biochemical approach to studying these problems in the nematode C. elegans. A set of monoclonal antibodies directed against germ-line-specific granules (P granules) has already been generated. In response to the asymmetry of the embryo, these granules are segregated specifically to the germ line or P lineage during early embryogenesis. P granules serve as an excellent marker for studying the mechanism of a symmetric distribution of cytoplasmic components to specific embryonic cells. Furthermore, they may act to specify germ-cell fates. The antibodies directed against P granules will be used in combination with molecular, cellular, and genetic techniques to address the following questions: 1) What is the nature of P granules? The antibodies will be utilized to purify P granules, identify their polypeptide components, and determine whether they contain nucleic acid. 2) Are P granules required for germ-cell development and, if so, how do they function? The antibodies will be used to specifically recover and characterize P-granule mutants. Antibodies will also be microinjected into living worms and embryos to try to functionally inactivate P granules. 3) How is P-granule gene expression regulated? The antibodies will be used to clone the genes encoding P-granule polypeptides, and the cloned sequences will be used to study the organization and transcription of the genes. 4) How are P granules segregated to the germ lineage? Because actin filaments appear to be involved in P-granule segregation, the organization of actin filaments appear to be involved in P-granule segregation, the organization of actin filaments in C. elegans embryos and the effects of actin inhibitors on granule segregation will be examined. The long range goal of the proposed experimental approach is to better understand how cell fates are specified early in embryonic development.

How the one-cell embryo generates the diverse array of tissues seen in adult organisms remains one of the central unsolved problems in development. In the nematode C. elegans, as in many species, this process is guided mainly by maternally supplied factors. My lab is taking advantage of the powerful genetics available in C. elegans to identify some of the genes that control this process. We have screened for mutations in genes that encode maternal components required for the specification and development of a specific cell type, the germ line. We have focussed on the germ line because it is not required for viability and mutant animals lacking germ cells are easily identified. We have isolated mutations that result in maternal-effect sterility or a "grandchildless" phenotype: homozygous mutant hermaphrodites produced by heterozygous mothers are themselves fertile but produce sterile progeny. Our screens have identified surprisingly few genes: six mes (for maternal- effect sterile) loci, defined by 23 mutations. The five characterized loci appear to belong to two classes. Embryos produced by mes-1 mutant mothers display defects in cytoplasmic partitioning during the division that generates the germ-line founder cell. The resulting larvae lack germ-line progenitor cells and contain extra body muscle cells. In contrast, the progeny of mes-2, mes-3, mes-4, and mes-6 mothers undergo normal embryogenesis, but show severe defects in post-embryonic proliferation of the germ line, resulting in agametic adults with 100-1000-fold reductions in germ cells. Our proposed experiments will address the following questions: Do mutations in mes-1 cause the germ-line founder cell to follow the fate of its sister, a muscle progenitor? Is this the null phenotype, or do more severe alleles affect earlier partitioning events and lead to embryonic lethality? Do the proliferation defects caused by mutations in mes-2, mes-3, mes-4, and mes-6 reflect defective determination of the germ line or an inability of the germ cells to execute their lineage? Are these mes gene products required in the germ line or in the somatic gonad? In addition to phenotype analysis, we will clone and molecularly analyze mes-1, mes-3, mes-4, and mes-6. Through immunolocalization of their gene products, we will learn whether any are partitioned to the germ line during early embryogenesis and whether any are germ-granule components. Finally, we will screen for and characterize mutations in additional mes loci. Our studies will elucidate the nature, localization, and function of maternal factors that participate in generating a functional germ line.

The normal program of early embryonic development is initiated when an oocyte is fertilized by a sperm. The mechanism by which sperm activate oocytes and the role of sperm components in this process and in later stages of embryogenesis are not understood. This project takes a combined genetic and molecular approach to studying this problem, using the nematode Caenorhabditis elegans as a model system. The PI has already identified a gene that encodes a sperm-supplied product that is required for initiation of the correct program of early embryonic development in C. elegans. Mutations in this gene, called spe-11, result in paternal-effect embryonic lethality: wild-type oocytes, when fertilized by sperm from mutant worms, undergo very abnormal early development and die. This is not due to the presence of a "poisonous product" in mutant sperm but to reduced levels or absence of wild-type product. Thus, wild-type spe-11 product participates, either directly or indirectly, in initiating the normal program of early development in C. elegans. This project has two major aims: 1) To clone and molecularly characterize the spe-11 gene, and localize the spe-11 gene product in sperm and newly fertilized embryos. 2) To carry out screens for additional paternal-effect embryonic-lethal mutations, which will identify other genes that encode sperm-supplied factors required for normal embryo development. This effort will complement the wealth of information on oocyte factors and will begin to define the sperm's contributions to early embryogenesis.

animal tissue; reproductive system; microscopy; model design /development; growth factor; biomedical resource;

The germ line enables metazoan organisms to produce gametes and thus is responsible for the fertility and perpetuation of species. We are investigating how the germ line is specified and its early development controlled, taking advantage of the powerful genetics available in the nematode worm C. elegans. The germ cells in many species contain distinctive granules, which may serve as determinants of germ-cell fate. We will elucidate the role of these germ-line "P granules" in C. elegans, by studying two genes that encode presumptive P-granule components. glh-1 was identified on the basis of its similarity to a Drosophila germ-granule component, vasa, and antibodies to GLH-1 stain P granules. pgl-1 was identified in genetic screens: pgl-1 mutant worms lack some P-granule epitopes and show a maternal-effect sterile phenotype (pgl/pgl mothers produce all sterile progeny). We will address the following questions: What is the mutant phenotype of glh-1? If glh-1 mutants are sterile (as predicted by anti- sense experiments), what are the germ-line defects? Is PGL-1 associated with P granules? Does PGL-1 resemble any known proteins? What are the germ-line defects in pgl-1 sterile worms? We will also use proven P- granule components (GLH-1 and perhaps PGL-1) to identify other P-granule components, for genetic and molecular analysis. We have identified four maternal-effect genes, mes-2, mes-3, mes-4, and mes-6, that are required for the normal early development and survival of the germ line. The maternal-effect sterile phenotype displayed by the four mes genes is more severe in XX than XO animals, suggesting that the mes genes participate in some aspect of control of gene expression that is sensitive to chromosome dosage. Consistent with this, MES-2 is similar to Drosophila Enhancer of zeste, which is predicted to control gene expression by regulating higher order chromatin structure. MES-6 is a WD- 40 repeat-containing protein, and MES-3 is a novel protein. We will address the following questions: Are the MES proteins localized to sites consistent with their being regulators of gene expression? Does MESA resemble any known proteins? Do the mes genes give a common mutant phenotype because they control each other's expression or localization, or because the MES proteins interact and function as a complex? Which of three models explains the sensitivity of the mes mutant phenotype to chromosome dosage? One model is that the mes genes control X chromosome dosage compensation in the germ line.

Research on the nematode worm Caenorhabditis elegans ranges from investigating such global problems as the circuitry and function of an entire nervous system and the organization of a genome, to investigating elements involved in control of gene expression and specification of the fates of individual cells. C. elegans has become an important experimental organism for the study of many aspects of animal biology, particularly the genetic and molecular bases of development and behavior. We are requesting funds to help cover costs and travel of participants to attend the International C. elegans Meeting to be held at the University of Wisconsin in Madison, WI, in June of 1999, and to be held either at the University of Wisconsin or at the University of California in Los Angeles, CA, in June of 2001. Previous C. elegans meetings have led to the exchange of knowledge, ideas, methods, mutants, and clones and have been vital in fostering the sense of excitement and the collegiality and cooperativity that characterize this field.

Research on the nematode worm Caenorhabditis elegans ranges from investigating such global problems as the circuitry and function of an entire nervous system and the organization of a genome, to investigating elements involved in control of gene expression and specification of the fates of individual cells. C. elegans has become an important experimental organism for the study of many aspects of animal biology, particularly the genetic and molecular bases of development and behavior. The biennial International C. elegans Meeting is a unique forum where researchers from the United States, Canada, Europe, and Asia can exchange the ideas and information that are essential for research on this organism. Plenary and poster sessions at the meeting present new findings in the areas of embryogenesis, cell fate determination, pattern formation, nuerobiology, cell biology, cell signaling and migration, cell death, sex determination, dosage compensation, the genome, and control of gene expression. In addition, several workshops provide opportunities for general discussion of new technologies, areas of investigation, and community efforts. Dr. Strome is requesting funds to help to cover costs and travel of participants to attend the International C. elegans Meeting to be held at the University of Wisconsin in Madison, WI, in June of 1999, and to be held either at the University of Wisconsin or at the University of California in Los Angeles, CA, in June 2001. Previous C. elegans meetings have led to the exchange of knowledge, ideas, methods, mutants, and clones and have been vital in fostering the sense of excitement and the collegiality and cooperativity that characterize this field.

DESCRIPTION (provided by applicant): Germ cells serve a unique role in animal development - they are not required for organism viability but are essential for fertility and thus for perpetuation of species. Special control mechanisms are required to ensure that germ cells survive from one generation to the next and are able to produce entire new organisms. We seek to understand how germ cells acquire and preserve these properties of immortality and totipotency. Our studies combine powerful genetic, genomic, and molecular approaches in the model system Caenorhabditis elegans. This proposal focuses on regulation of chromatin states and gene expression patterns in germ cells. We previously identified four C. elegans MES proteins as being required for germ cell survival and fertility and showed that their major role is silencing the X chromosomes. MES-2, MES-3, and MES-6 function as a complex to concentrate a repressive histone modification, methylation of histone H3 on Lys27, on the Xs. In contrast, MES-4 is dramatically concentrated on the autosomes, where it methylates histone H3 on Lys36. Despite its apparent absence from the X, removal of MES-4 leads primarily to desilencing of genes on the X. Our working model for X silencing is that MES-2/3/6 action directly represses gene expression and that MES-4 acts at a distance, by repelling repressors from autosomal regions and focusing their action on the Xs. The aims of this proposal are to: 1) Test that model and identify new participants in MES regulation by investigating candidate genes and using a powerful unbiased genetic screen. 2) Learn at the gene level where MES-4 and its H3Lys36 methyl marks are located and investigate how MES-4 is recruited to those sites and how the MES-2/3/6 complex keeps MES-4 off the X chromosomes. These studies will use chromatin immunoprecipitation followed by tiling array (ChIP-chip) approaches. 3) Gain a high resolution view of the distribution of MES-2/3/6 and its H3Lys27 methyl marks across the genome and test whether autosomal MES-4 participates in concentrating MES-2/3/6 on the Xs. This aim will also take advantage of ChIP-chip technology. In addition to insights into developmental strategies used in C. elegans germ cells, our studies will shed light on the diverse roles and regulation of H3Lys27 and H3Lys36 methylation across species, how cells regulate the chromatin state of large domains and entire chromosomes, for instance during dosage compensation, and how mammalian homologs of the MES proteins contribute to stem cell biology and cancer.

Developmental biology is one of the most exciting areas of modern biology. This award will help fund the 2008 Santa Cruz Developmental Biology (SCDB) Meeting. SCDB meetings are international in scope but are also highly accessible to junior faculty, postdoctoral fellows, and graduate students. The 2008 Meeting will be organized around the theme "Transitions in Development". The meeting will have 49 platform speakers, of which 21 will be chosen from abstracts submitted to the meeting organizers, allowing postdocs and graduate students ample opportunity to present their own data. Two poster sessions allow all other attendees to present their work. The 2008 SCDB Meeting will have a significant positive impact on the field of developmental biology, educating the ~200 participants in the latest advances in this exciting area, disseminating the results in a meeting report in a leading journal in the field, and fostering training for scientists in the early stages of their careers. Efforts will be made to attract a group of participants that represents the diversity of the field in organism studied and approach used that is balanced in gender, and that increases participation by underrepresented minorities. This award will help achieve this goal, by providing meeting associated registration and housing costs for eight graduate students and postdoctoral fellows, selected to increase the diversity of the participants, as well as registration and housing costs for four junior faculty members who will present talks.

Developmental biology has a central place in the life sciences. By focusing on the study of whole organisms using a wide variety of approaches and technologies, the field has contributed significantly to our understanding of the common processes that regulate the development of life on our planet. Over the last decade, developmental biologists have made significant, Nobel-prize winning, discoveries in aging and in the control of cell death (Nobel 2002). They have worked out the mechanisms by which small RNAs turn genes on or off (Nobel 2006 and Lasker award in 2008) and have driven the development of new techniques; for example gene replacement in mammals (Nobel 2007) and the ability to see proteins in living cells (Nobel 2008). The 2010 Santa Cruz Developmental Biology (SCDB) meeting provides a forum for developmental biologists working at a variety of levels and using a wide range of experimental subjects to come together under the theme of 'Diverse strategies in Development'. Here, researchers can learn about the newest major breakthroughs and how these can further accelerate their progress in fields under the umbrella of developmental biology. Over 150 developmental biologists are expected to attend the SCDB meeting at University of California, Santa Cruz from June 30th--July 3rd, 2010 to discuss recent progress, to exchange ideas and to educate both the current leaders in the field and upcoming researchers and students. Due to the high visibility and quality of the invited speakers participating in this endeavor and the small size of the meeting, the SCDB provides unprecedented opportunities for interactions between established and young scientists. The conference program is well balanced to promote participation of women and members of groups traditionally underrepresented in science.

DESCRIPTION (Provided by Applicant): The investigators are organizing the Santa Cruz Developmental Biology (SCDB) Meetings around a theme, "Diverse Strategies in Development". The goals of the SCDB Meetings are to educate the community about the latest major advances in developmental biology through the dissemination and discussion of current data and emerging concepts. Simultaneously, the meetings facilitate scientific networking among developmental biologists at all stages of their careers. This is possible because the meeting size is relatively small and fosters interactions, yet the caliber of science is exceptionally high. The organizers also bring together a diverse group of scientists who use genomic, genetic, biochemical, molecular, cellular, and classical embryological approaches to bear on the most important aspects of development across the full range of plant, animal, and microbial model systems. This assembly is a single-platform style meeting occurring over a period of one evening and three full days, and is comprised of seven sessions. Each session consists of four 25-minute invited talks and three 15-minute talks selected from the abstracts. All 28 of the invited speakers gave enthusiastic, affirmative responses upon receiving invitations to participate. Seven of them are session chairs and have been recruited from the ranks of the leaders in their fields. In order to provide the opportunity for the widest range of speakers, the organizers will choose an additional 21 platform presentations from the abstracts. Speakers selected from abstracts will preferentially be students, postdoctoral fellows, and early stage investigators, and the organizers aim for gender parity. PROJECT NARRATIVE: Research in developmental biology has contributed significantly to our understanding of the fields of congenital diseases, stem cell biology, oncology, and regenerative medicine. Understanding the cellular and molecular control of developmental processes will help us decipher the causes of birth defects, and ultimately lead to new techniques for diagnosis and treatment. In the 2010 SCDB meetings, the organizers will learn how various fields are making major breakthroughs and how these can further accelerate the progress in science in fields under the umbrella of developmental biology

DESCRIPTION (provided by applicant): The UCSC graduate training program in Molecular, Cellular, and Developmental Biology provides intensive training in the skills necessary for outstanding research in modern biology. First year trainees undertake coursework emphasizing critical evaluation of scientific models and experimental results, and they also participate in three ten-week laboratory rotations. Second-year students take an oral qualifying exam, and advanced students participate in a variety of seminars, advanced special topics courses, and research group meetings designed to provide continuing learning opportunities. The goal of the training program is to produce graduates who have a strong foundation in the specific area of their thesis research, as well as broad training and knowledge in molecular, cellular, and developmental biology. The program is administered by Advising and Admissions Committees according to policy guidelines stated in our Graduate Handbook and Faculty Guidelines. The training program is composed of 19 members of the Department of IVICD Biology, and 9 affiliated faculty from the Departments of Biomolecular Engineering, Chemistry and Biochemistry, and Microbiology and Environmental Toxicology. The majority of the faculty members are housed in the Sinsheimer Laboratories, with other faculty located in the Physical Sciences Building, which is adjacent to Sinsheimer Labs. The MCD training program provides an interactive and collaborative research environment for graduate training. Research in the training program utilizes a wide variety of approaches and model organisms, and is organized as interdisciplinary clusters of faculty with shared interests, thereby creating critical masses of researchers that foster mutual support and scientific interaction. During the first 10 years of NIH training support we have recruited and trained outstanding students who would be amongst the best students in any training program. We are requesting a total of 8 predoctoral trainee positions in this renewal, which will continue to be awarded to our best students. Relevance: Students in the MCD Biology training program have extensive opportunities to study topics related to human health. Many training faculty work on systems that are directly relevant to human health, including stem cell biology, malaria, cholera, cancer biology, host-pathogen interactions, neurodegenerative diseases, and the responses of neurons to stroke and other damage. Other faculty carry out basic research that provides the foundation for understanding topics relevant to human health, including research focused on cell division, signaling, meiosis, chromatin remodeling, cell differentiation, and neurobiology.

Nematodes (roundworms) are the most abundant multicellular animals on earth, colonizing virtually every terrestrial, marine and freshwater habitat. While the nematode laboratory model C. elegans is the focus of intense, technically sophisticated research, much less is known about other nematodes, despite their biological importance. Of particular significance are parasitic nematodes, which comprise more than half of the estimated 28,000 nematode species; these parasites have important effects on the biology of their plant and animal hosts, threatening agricultural crops, and causing debilitating human diseases that affect over a billion people worldwide. By promoting interactions and collaborations between C. elegans researchers and investigators of parasitic nematodes, this project will increase awareness among C. elegans scientists of the issues and problems that parasitic nematode researchers face, and will pave the way for application of the powerful molecular and cell techniques developed in C. elegans research to investigations of parasitic nematodes. The planned symposium at the International C. elegans Meeting (Los Angeles June 26-30th, 2013) will provide an exceptional opportunity to reach the majority of C. elegans researchers. Six internationally recognized experts on plant, animal and human parasitic nematodes will speak on the life history and unique biology of parasitic species and on the outstanding issues in their field. In addition to formal education of the C. elegans community, this session will provide time for researchers in the two communities to interact more informally as well. The symposium aims to stimulate C. elegans researchers to think about problems facing the study and control of parasitic nematodes and to apply their expertise toward solving these problems. This session will be particularly valuable for undergraduate and graduate students and post-docs by exposing them to new possible career paths. Funds will also be provided for under-represented minority graduate students to attend the session.

DESCRIPTION (provided by applicant): The goals of the UCSC Graduate Training Program in Molecular, Cell and Developmental Biology are to recruit highly motivated graduate students, provide them with critical thinking skills, expertise in experimental design and interpretation, an a strong framework of knowledge, while also amplifying their passion for inquiry and discovery. We strongly encourage interdisciplinary and collaborative research that crosses departmental, divisional, and even institutional boundaries. Intellectual, cultural, and socioeconomic diversity are prevalent in our graduate student population, and we constantly seek new approaches to attract and inspire students from under-represented groups. Our Training Program prepares students to pursue diverse careers in biomedical research, teaching, government, and industry. First-year trainees participate in three 6-week research rotations and in a core curriculum designed to train them in the logic of experimental analysis. Students learn how to evaluate published research, effectively argue scientific points, formulate hypotheses, design rigorous experiments, and write research manuscripts and proposals with intellectual depth. Students join a laboratory to pursue their Ph.D. thesis research in Spring of their first year. Second-year students train in the responsible conduct of research and take an oral qualifying exam. Third-year students present a full research seminar to the Training Program. Students at all stages participate in departmental seminars, research meetings, and journal clubs, and take advanced elective courses of their choosing. These broaden their training experience and promote interactions among trainees and faculty. Training culminates with a written Ph.D. dissertation and oral defense. Our Training Program includes 30 faculty from 5 departments, including the Departments of Molecular, Cell and Developmental Biology, Biomolecular Engineering, Chemistry and Biochemistry, Microbiology and Environmental Toxicology, and Physics. Participating laboratories are intermingled in 3 adjacent buildings on science hill, which encourages interaction and collaboration. Research in the Training Program utilizes a wide variety of approaches, and is organized with interdisciplinary clusters of common interests (e.g. Cell Biology, Chromatin, Neurobiology, and RNA), thereby creating critical masses of researchers that foster mutual support and scientific creativity. Many training faculty work on systems that are directly relevant to human health, including stem cell biology, malaria, cholera, cancer biology, host-pathogen interactions, neurodegenerative diseases, and the responses of neurons to stroke and other damage. Other faculty carry out basic research that provides the foundation for understanding topics relevant to human health, including cell division, signaling, meiosis, chromatin organization, cell differentiation, and neurobiology. Our Training Grant, currently in its 14th year, supports 6 trainees. In this renewal, we are requesting support for 8 trainees. Training Grant positions will continue to be awarded to our most promising students.

DESCRIPTION (provided by applicant): The goals of the UCSC Graduate Training Program in Molecular, Cell and Developmental Biology are to recruit highly motivated graduate students, provide them with critical thinking skills, expertise in experimental design and interpretation, an a strong framework of knowledge, while also amplifying their passion for inquiry and discovery. We strongly encourage interdisciplinary and collaborative research that crosses departmental, divisional, and even institutional boundaries. Intellectual, cultural, and socioeconomic diversity are prevalent in our graduate student population, and we constantly seek new approaches to attract and inspire students from under-represented groups. Our Training Program prepares students to pursue diverse careers in biomedical research, teaching, government, and industry. First-year trainees participate in three 6-week research rotations and in a core curriculum designed to train them in the logic of experimental analysis. Students learn how to evaluate published research, effectively argue scientific points, formulate hypotheses, design rigorous experiments, and write research manuscripts and proposals with intellectual depth. Students join a laboratory to pursue their Ph.D. thesis research in Spring of their first year. Second-year students train in the responsible conduct of research and take an oral qualifying exam. Third-year students present a full research seminar to the Training Program. Students at all stages participate in departmental seminars, research meetings, and journal clubs, and take advanced elective courses of their choosing. These broaden their training experience and promote interactions among trainees and faculty. Training culminates with a written Ph.D. dissertation and oral defense. Our Training Program includes 30 faculty from 5 departments, including the Departments of Molecular, Cell and Developmental Biology, Biomolecular Engineering, Chemistry and Biochemistry, Microbiology and Environmental Toxicology, and Physics. Participating laboratories are intermingled in 3 adjacent buildings on science hill, which encourages interaction and collaboration. Research in the Training Program utilizes a wide variety of approaches, and is organized with interdisciplinary clusters of common interests (e.g. Cell Biology, Chromatin, Neurobiology, and RNA), thereby creating critical masses of researchers that foster mutual support and scientific creativity. Many training faculty work on systems that are directly relevant to human health, including stem cell biology, malaria, cholera, cancer biology, host-pathogen interactions, neurodegenerative diseases, and the responses of neurons to stroke and other damage. Other faculty carry out basic research that provides the foundation for understanding topics relevant to human health, including cell division, signaling, meiosis, chromatin organization, cell differentiation, and neurobiology. Our Training Grant, currently in its 14th year, supports 6 trainees. In this renewal, we are requesting support for 8 trainees. Training Grant positions will continue to be awarded to our most promising students.