REGULATION OF DIGESTION IN BLOOD-SUCKING INSECTS

Blood-sucking insects transmit a number of important diseases. Control
programs based on the use of insecticides had early success, but the recent
development of resistant insects has made such measures problematic. It is
now clear that control measures based on the biology of the insect, so-
called biorational control, should be developed. Relatively little work
using modern biochemical and molecular biological approaches has been done
on insect vectors. The ingestion of the blood meal induces many
morphological and biochemical changes in the midgut, which are required for
digestion of the blood meal. The long-range goal of this research is to
understand this differentiation of the midgut and to identify potential
steps in this process which might serve as targets for controlling these
insects. In this application we propose to use two important disease
vectors: Aedes aegypti, a mosquito which transmits yellow fever and dengue
fever, and Rhodinus prolixus, a hemipteran which transmits Chagas disease.
We propose to study how the blood meal regulates the expression of the
genes encoding the major midgut digestive proteases as a paradigm for
understanding the differentiation of the midgut. A. aegypti uses a
trypsin-based digestive system, whereas R. prolixus uses a cathepsin-based
digestive system. In A. aegypti there is evidence for the existence of two
forms of trypsin, an "early" form, which appears immediately after the
blood meal, and a "late" form, which is responsible for most of the
digestion of the protein in the blood meal. In R. prolixus digestion in
the lumen of the midgut is carried out by enzymes which are normally found
in lysosomes. The specific aims of this proposal are as follows: 1) clone
and sequence the cDNA for the "early" trypsin from the midgut of the
mosquito Aedes aegypti; 2) clone and sequence the genes for the "early" and
"late" trypsin from Aedes aegypti; 3) clone and sequence the cDNAs and
genes for the cathepsin B and D from the midgut of Rhodinus prolixus; 4)
Use cDNA and antibody probes to analyze factors in the blood meal, or
factors produced in the insects in response to the blood meal, which affect
the expression of the genes encoding the digestive proteases; 5) Identify
cis-acting elements in the protease genes and use these DNA sequences to
isolate cDNAs for the trans-acting factors which bind to the cis-acting
elements. Sequence these cDNAs and use them to investigate how the blood
meal affects the amount or activity of the trans-acting factors.