Henry H. Hagedorn - US grants

Dr. John Law and his colleagues at the University of Arizona will establish a Biological Research Center for Insect Science (CIS). The CIS will aim research at 1) discovering new aspects of insect biology that will aid in the design of biorational methods for controlling insects; 2) development of model systems using insects that will yield fundamental knowledge useful in the study of mammalian systems; and 3) searching for new useful products and processes from insects. Programs will include research, training, education and outreach, with a strong emphasis on molecularly-oriented interdisciplinary research. The CIS will sponsor meetings and symposia in insect science, and have strong international linkages. An industrial associates program will facilitate technology transfer. Individual projects will include research on insect: cuticle structure and function, digestion and nutrition, proteins and hormones, endocrine systems, reproductive systems and central nervous systems. Other studies will focus on molecular, sensory, behavioral and ecological factors in insect-plant interactions; insect communication and behavior; insecticide action; caste differentiation in social insects; and insect population biology, systematics and evolution. Through the results of its research and training programs, the CIS should have a worldwide impact on fundamental and applied biology. Insects share co-dominance of the biosphere with humans. The relationship ranges from competitive to cooperative, as they compete for resources, carry disease, pollinate plants, and process organic wastes. The importance of understanding insect biology was brought out as some chemical pesticides were abandoned for social and environmental concerns, and as the effectiveness of others decreased when resistant strains developed. In fact, much of our knowledge of insects derives from studies of one organism, the common fruit fly Drosophila, which has been exploited for decades in the study of genetics and animal development. The remarkable new experimental tools provided by recent advances in biochemistry and molecular biology now allow new approaches to the study of fundamental aspects of the biology of many other insects. Besides contributing to the development of new agents and strategies for pest control, such studies should provide a wealth of information on genetics, social behavior, animal development, and nervous system function. To this end, the CIS will provide multidisciplinary research and education programs on insect science. This center will be built on a strong base of existing faculty and will coordinate with an existing Biotechnology Center to expand collaborative efforts in basic research, to train present and future scientists, and, in collaboration with industry, to probe the potential uses of insects in applied research.

In sheer numbers, insects are the most successful species on earth. No one denies the many beneficial activities of insects as, for example, in the pollination of economically important crops, the production of silk, and most recently biological pest control (one insect predating on another). Everybody also knows about the potentially devastating effects of insects. Insects have been known to wipe out entire crops, contributing to world hunger. Insects are also known as the dreadful bearers of blood borne fatal diseases such as yellow fever, malaria, and elephantiasis. Since biblical time (the flight of locust) men have tried to cope with the undesirable impacts of insects on mankind. In modern times the most successful approach has been the use of pesticides; but pesticides - as we are now learning - have serious undesirable side-effects on man and animal alike. For this reason alternatives to insect control are needed. The present proposal marks the exploration of a new approach via attempts to interfere in the normal salt and water budget of the insect. This approach will be pursued in the blood-feeding yellow-fever mosquito (Aedes aegypti). Shortly after mating the female mosquito seeks a bloodmeal. She needs blood in order to obtain nutrients and proteins for her eggs. Having found a source, the female yellow-fever mosquito gorges herself, taking a meal 2-3 times her own body weight| Such a heavy payload prevents the mosquito from flying away, thereby threatening reproductive success (she must find a pond to deposit her eggs) and personal calamity (a swat). To guarantee the former and to avoid the latter, she must maintain flight capability and maneuverability, namely by quickly eliminating unwanted weight, in particular the unwanted salt and water portion of the bloodmeal. This she does by way of a hormone, the so-called diuretic, natriuretic hormone. In the upcoming grant period the PIs plan to purify and isolate this hormone, learn its structure and attempt to synthesize it. This work will be done in the laboratory of a peptide chemist, Dr. Hayes of Texas A&M University. At the same time Dr. Beyenbach (a transport physiologist) at Cornell University will study how the kidney in the mosquito works normally and under the influence of the hormone. And the laboratory of the insect endocrinologist, Dr. Hagedorn (University of Arizona) will investigate how the hormone is released in the mosquito, where it is made and stored, and what organs other than the kidney it affects. Three potential target sites for pest control via disrupting normal salt and water balance in the insect will be investigated: 1) by giving synthetic hormone causing the insect to lose salt and water when they need to be conserved, 2) by interfering with normal kidney function, e.g. preventing the action of the hormone and thereby reproductive success, and 3) by disrupting the normal feedback loop of salt and water balance by inhibiting the production or the release of the hormone, when the insects need the hormone.

This work will contribute to an understanding of the reproductive physiology of the mosquito, an important vector of human and animal diseases including malaria, yellow fever, dengue, encephalitis and others worldwide. The advent of recombinant DNA technology has raised the hope that the mosquito could be genetically engineered, with the aim of preventing transmission of disease-causing organisms. It is, therefore, vital that we understand how genes are regulated in this insect. This work will focus on the control of gene expression at the molecular level in a mosquito. A long term goal of the research is to contribute to the development of technology that will allow the genetic manipulation of mosquitoes. Genes that play a role in mosquito reproduction provide an essential tool for developing the molecular biology and genetics of this important pest insect. The present proposal is focused on female-specific genes that are expressed during egg development in the response to the steroid hormone, 20-hydroxyecdysone. We plan to build upon earlier studies in which we have identified, from genomic libraries, 4 clones encoding vitellogenin genes - which are expressed only in the fat body in response to ecdysone - and another clone, containing an ecdysone-inducible gene expressed specifically in ovarian follicle cells. An important goal of this proposal is to characterize the 5'- regulatory sequences of these genes, with the ultimate goal of developing constructs that can be used to define nucleotide regions that confer ecdysone responsiveness and tissue-specificity of gene expression. Since functional analysis of gene expression in mosquitoes requires efficient transformation techniques, the development of which has been severely hampered by the lack of cloned mosquito genes, a second major goal of this proposal is to optimize a mosquito transfection system to allow the rapid analysis of hormone-inducible gene constructs in vitro. It is clear that genetic transformation of mosquitoes is a reasonable goal given the powerful tools made available by recombinant DNA technology, and the rapid advances being made in the transformation of a wide variety of plants and animals. Given the obvious medical importance of the mosquito, the choice of this insect as a particular target for genetic transformation is apparent.

The goal of this proposal is to examine the molecular basis of the physiological phenomenon termed `competence' which is used here to describe the appearance of responsiveness to a hormone that results in the expression of new genes. The appearance of the competent state is thought to be due to structural changes in regulatory regions of these genes that permit the binding of proteins, such as hormone receptors and other transcriptional factors, to specific nucleotide sequences. This study should help us to understand how hormones regulate gene expression. Two genes expressed during egg development in the mosquito, Aedes aegypti, will be examined; a vitellogenin and a vitelline envelope protein gene, both of which are under the control of 20- hydroxyecdysone. We have shown that the vitellogenin gene of the newly enclosed adult female is not capable of responding to 20- hydroxyecdysone, and that competence to respond appears during the first two days after eclosion in response to juvenile hormone. We propose to use a mobility shift assay to follow changes in the binding of proteins to the regulatory regions of these two genes during the acquisition of competence and after exposure to 20- hydroxyecdysone and juvenile hormone.

molecular biology; meeting /conference /symposium; Insecta; travel;

DESCRIPTION (Adapted from the applicant's abstract): Using insects as tools, the investigators on this application will design an active-learning program that will increase students' scientific literacy and reasoning skills. The instructional materials will be multi-level and multidisciplinary, providing many links between science and everyday life, and will enable teachers to draw in students of many levels and cultural backgrounds. Previous results have clearly demonstrated that the presence of live insects in classrooms generates excitement, focuses attention, and improves retention of lesson content. The increasing failure of insecticides to control insects is linked to a consideration of toxicology and biological control. The overwhelming importance of the most decimating and prevalent human diseases (e.g., malaria) and food and fiber crops in all cultures provide innumerable links to human health, history, economics, and culture throughout the world. Instructional materials will be designed to exploit these connections to heighten students' interest in health sciences. Students will meet active scientists through video presentations and share data, analyses, and experiences with scientists and other classrooms through the Internet. The instructional program will be created through a partnership with teachers, piloted in local schools and tested at distant sites to validate its effectiveness in other regions.