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Regulation of tissue and development specific gene expression patterns underlies the functional specialization of organs in multi-cellular organisms. In the viviparous tsetse fly ( Glossina ), the female accessory gland is specialized to generate nutrients in the form of a milk-like secretion to support growth of intrauterine larva. Multiple milk protein genes are expressed specifically in the female accessory gland and are tightly linked with larval development. Disruption of milk protein synthesis deprives developing larvae of nutrients and results in extended larval development and/or in abortion. The ability to cause such a disruption could be utilized as a tsetse control strategy. Here we identify and delineate the regulatory sequence of a major milk protein gene ( milk gland protein  1: mgp1 ) by utilizing a combination of molecular techniques in tsetse,  Drosophila  transgenics, transcriptomics and  in silico  sequence analyses. The function of this promoter is conserved between tsetse and  Drosophila . In transgenic  Drosophila  the  mgp1  promoter directs reporter gene expression in a tissue and stage specific manner orthologous to that of  Glossina . Analysis of the minimal required regulatory region of  mgp1 , and the regulatory regions of other  Glossina  milk proteins identified putative homeodomain protein binding sites as the sole common feature. Annotation and expression analysis of  Glossina  homeodomain proteins identified  ladybird late  ( lbl ) as being accessory gland/fat body specific and differentially expressed between lactating/non-lactating flies. Knockdown of  lbl  in tsetse resulted in a significant reduction in transcript abundance of multiple milk protein genes and in a significant loss of fecundity. The role of Lbl in adult reproductive physiology is previously unknown. These results suggest that Lbl is part of a conserved reproductive regulatory system that could have implications beyond tsetse to other vector insects such as mosquitoes. This system is critical for tsetse fecundity and provides a potential target for development of a reproductive inhibitor.

The Homeodomain Protein Ladybird Late Regulates Synthesis of Milk Proteins during Pregnancy in the Tsetse Fly (Glossina morsitans)