Myo-inositol is a six-carbon sugar alcohol. It is essential as a precursor of the phospholipid membrane component phosphatidylinositol (PI) and the phosphoinositide signaling pathway in all eukaryotes. It aids in cellular metabolism, osmoregulation, and plays an important role in fertilization and diseases such as diabetes, bipolar disorder, and Alzheimer’s disease. Myo-inositol metabolism is comprised of synthesis, transport, catabolism, and recycling. Myo-inositol synthesis is catalyzed by myo-inositol-3-phosphate synthase (MIPS). Surprisingly, synthesis of myo-inositol and its role in fertilization has not yet been studied in the model organism Drosophila melanogaster (fruit fly). We hypothesize that MIPS expression is essential for growth and development of D. melanogaster. In this study, a precise deletion of the entire MIPS gene was generated and confirmed through PCR amplification and sequencing of the resultant DNA fragments. The lack of the MIPS transcript in homozygous MIPS deletion flies was confirmed by RT-PCR. During that experiment, two additional isoforms of MIPS were identified in wild-type flies (CS). Supplementation of chemically defined food with 0.5mM inositol was required to sustain all homozygous MIPS deletion fly strains. Fully-grown homozygous deletion flies could live without additional inositol in the food, but newly emerged larvae only survived to the first instar larval stage. However, even while on rich media supplemented with 170mM inositol, a homozygous MIPS deletion stock was unable to produce viable offspring. Homozygous MIPS deletion strains were identified as male-sterile, incapable of producing offspring when mated to any strain of females (including wild-type). Homozygous female MIPS deletion flies were fertile and maintained a high fecundity rate when mated to any strain (with an exception of homozygous male MIPS deletion flies). The male-sterility was complemented with the addition of a wild-type MIPS gene to chromosome 3. Testes dissections of homozygous male MIPS deletion flies revealed improper progression of spermatogenesis, specifically during sperm individualization. These studies contribute to the understanding of the role of inositol synthesis in growth, development, and fertilization.
|Advisor:||Klig, Lisa S.|
|Commitee:||Brusslan, Judy, Eldon, Elizabeth|
|School:||California State University, Long Beach|
|School Location:||United States -- California|
|Source:||MAI 55/02M(E), Masters Abstracts International|
|Subjects:||Molecular biology, Developmental biology|
|Keywords:||Drosophila melanogaster, Fertilization, Myo-inositol, Myo-inositol-3-phosphate synthase|
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