Vivipararous reproduction is a relatively uncommon process in insects. Tsetse flies (family Glossinidae) are one of only a few families of flies (Hippoboscidae, Nycteribiidae, Calliphoridae and Streblidae) that are capable of undergoing pseudo-placental viviparous reproduction (intrauterine larval development and nourishment) (Meier et al., 1999
). This strategy results in fewer offspring per female, but a higher level of survival and fitness for the offspring. In order to accommodate this lifestyle, tsetse has undergone radical changes in its reproductive physiology and regulation of reproductive processes.
Oogenesis in tsetse is a process that has undergone major changes in relation to how it occurs in other insects. In Drosophila
, a close relative of tsetse, oogenesis occurs asynchronously in multiple ovarian follicles in both ovaries (Bownes, 1982
). Vitellogenesis in Drosophila
occurs by the synthesis and secretion of yolk proteins from both the fat body and follicle cells of the ovaries. However, vitellogenesis in tsetse has been reduced in scale to suit their reproductive cycle. They have a small number of ovarioles per ovary (two) and only develop a single oocyte at a time. Oocyte development occurs in alternating ovaries. The first oocyte to be developed can always be found in the right ovary (Denlinger and Ma, 1974
). These changes have significant implications for the regulation of genes associated with oogenesis, such as GmmYP1
. Expression of GmmYP1
appears to be restricted to the reproductive tract and is under strict physiological regulation. The fact that only one oocyte develops at a time implies that expression of this gene may only occur in a subset of cells within the ovaries (most likely within the ovariole carrying the currently developing oocyte). The signals that are determining which follicles will develop and which will wait are unknown. The sequential nature of how the active follicles change between the left and right ovaries implies a mechanism of communication between the ovaries and their follicles as to their oogenic status. This mechanism may regulate the expression of GmmYP1
so that it occurs in the appropriate oocyte at the correct time. The presence of a developing embryo or larvae within the uterus also appears to have a regulatory effect upon GmmYP1
transcription. When a developing larva is present in the uterus and an oocyte is waiting to ovulate, levels of GmmYP1
transcript are minimal. Once the larva has been deposited and the next oocyte is ovulated into the uterus, GmmYP1
transcription increases again in concordance with the next round of oogenesis. The mechanisms associated with this regulatory system are not yet understood and will be the target of future research. Protein levels for GmmYP1 appear to be fairly stable after development of the initial oocyte. This is not unexpected, as different stages of oocyte are present at almost all the stages of the reproductive cycle.
The most dramatic modification of tsetse reproductive physiology is the evolution of physiology capable of sustaining an offspring throughout the duration of its larval development. A key component to this physiology is the accessory glands (milk glands), an organ consisting of tubules that intertwine throughout the fat body in the abdominal cavity and coalesce at an opening to the uterus where the developing larva feeds. This gland is lined with cells that generate large amounts of protein that is secreted along ducts running through the center of the tubules. However, tsetse is not the only insect known to produce nutrients for its offspring in the form of milk. The viviparous cockroach, Diplotera punctata
, also produces milk to nourish its developing embryos (Stay and Coop, 1974
). The milk is generated from secretory cells that line the walls of the brood sac where the embryos undergo development. Analysis of protein from the cockroach milk has revealed that it consists mainly of a molecule in the lipocalin family, the same family from which GmmMGP originates (Williford et al., 2004
). This is an interesting observation, as proteins from the lipocalin family have also been associated with lactation in marsupials (Piotte et al., 1998
). The occurrence of lipocalins in lactation products of divergent organisms suggests that lipocalins have evolved to fill the role of milk proteins multiple times. A common property of lipocalins is the ability to carry small hydrophobic molecules. These observations suggest that GmmMGP, besides being a source of raw amino acids, may also be involved in the transport of hydrophobic substances from mother to offspring.
Regulation of GmmMGP
appears to be both transcriptional and translational. Transcripts for GmmMGP
are undetectable until the first larva hatches in the uterus. This suggests that expression of the gene might be dependent upon signals produced by the larva, or the pregnancy state of the mother. Once transcription of GmmMGP
begins, its expression is relatively constant. However, fluctuation of protein levels at the time of partuition suggests the possibility of translational control as well. The low levels of GmmMGP immediately after birth suggests that its translation may cease until embryonic development of the next offspring has been completed and the next larva hatches into the uterus. Previous work suggests that milk production may be regulated via juvenile hormone, as the corpora allata (CA) (the gland responsible for juvenile hormone synthesis) undergoes cyclical changes in volume and histological appearance in parallel with the pregnancy cycle (Ejezie and Davey, 1974
). Furthermore, ablation of the CA reduces milk synthesis in tsetse flies. The phenotype caused by CA ablation is reversible by ectopic treatment of flies with juvenile hormone (Ejezie and Davey, 1976
). Further analyses of GmmMGP
transcriptional and translational regulation and the role of juvenile hormone in this system will be the subject of future research.
The results obtained in this work form a molecular foundation on which a functional analysis of tsetse reproductive function can be developed. These data confirm the association of the GmmYP1 and GmmMGP genes with oogenesis and larvigenesis processes, respectively. Therefore, it is possible to use the dynamics of these proteins as markers of reproductive function in experimentally compromised flies. Future work in this area will focus on the identification of signals that regulate the transcriptional and translational processes of these genes. Moreover, future experiments will include the determination of the effects of various physiological states, such as nutritional deprivation, aposymbiosis, trypanosome infection and absence of mating stimulus on reproductive gene expression. These experiments will elucidate the regulatory mechanisms controlling reproduction in tsetse, and possibly reveal new strategies that can be applied to tsetse population control.