An interspecific functional complementation test in Drosophila for introductory genetics laboratory courses.
Introductory genetics courses often include evolutionary genetics concepts such as sequence homology and functional conservation. It is usually assumed that two sequences showing homology (i.e., sharing a common ancestral sequence) perform the same molecular function. The correlation, however, does not always hold true, and evidence for functional conservation must come from functional studies. In this study we describe a genetics laboratory class that demonstrates functional conservation between the Drosophila protein Muscleblind (Mbl) and its human ortholog MBNL1. We use the Gal4/UAS system to express MBNL1 in a Drosophila mutant background and measure the in vivo activity of the human protein by rescue of mbl mutant phenotype in embryos. As a control, ubiquitous expression of Drosophila MblC, one of the four protein isoforms encoded by the gene, increased by 71% the viability of mbl mutant embryos and greatly reduced the hypercontracted abdomen of mutant larvae. In a parallel experiment, human MBNL1 provided a robust rescue of the embryonic lethality (78%) and improved abdomen hypercontraction as well. Under both conditions, rescued larvae die as first instars, probably due to overexpression effects, lack of alternative protein isoforms, or incomplete expression in critical tissues such as the nervous system. The use of two constructs in the rescue experiment (UAS-mblC and UAS-MBNL1) and the incomplete rescue prompt several questions for students. The fact that a human protein works in a Drosophila cellular context illustrates the use of an in vivo test to prove functional conservation.
Monferrer
Lidon
L
Department of Genetics, Valencia University, Doctor Moliner 50, Burjasot 46100, Valencia, Spain.
Artero
Ruben
R
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D004330:Drosophila / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D005816:Genetic Complementation Test / Q000379:methods*; D005823:Genetics / Q000193:education*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D026801:Synteny
D029721:Drosophila Proteins; C493663:MBNL1 protein, human; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; C109271:mbl protein, Drosophila