Comprehensive characterization of the complex lola locus in Drosophila melanogaster reveals novel roles in vivo
Date issued
Authors
Editors
Journal Title
Journal ISSN
Volume Title
Publisher
License
Abstract
longitudinals lacking (lola) is one of most complex genes in Drosophila, encoding for 20 different protein isoforms by alternative cis- and trans-splicing. Each splice variant shares a constitutive N-terminus but contains distinct isoform-specific C-terminal exons, which for most isoforms encode a zinc finger motif. Lola acts as a transcription factor playing diverse roles in vivo, including axonal guidance and neuronal maintenance. Majority of previously performed studies employed loss-of-function alleles disrupting all Lola isoforms, making it difficult to decipher isoform-specific roles during development. Therefore, the physiological function of individual Lola isoforms has been poorly characterized.
My PhD project aimed to better characterize physiological functions of Lola isoforms in vivo. Using the recently developed CRISPR/Cas9 system, I created mutations in isoform-specific C-terminal exons and mutant alleles were generated for all 20 Lola isoforms. Phenotypic analysis revealed an opposing function for the isoforms Lola-A and Lola-H in regulating locomotion behaviour of adult flies. Furthermore, a critical role was identified for Lola-O in controlling octopamine synthesis and thus preventing neurodegeneration. This splice variant is specifically expressed in octopaminergic neurons, where it regulates the expression of the Tyramine-ß- hydroxylase, a key enzymatic component of the octopamine synthesis pathway. Lastly, I identified Lola-F as an essential factor controlling axonal pathfinding along the embryonic ventral midline. Lola-F positively regulates the expression of several axon guidance genes, including the microtubule-associated factor Futsch.
This study emphasizes the significance of the CRISPR/Cas9 system in sequence specific genome modification and its concomitant importance on the functional analysis of distinct protein isoforms. Furthermore, this work demonstrates that Lola splice variants exhibit a high functional diversity in distinct cell types, thereby underlining the importance of alternative splicing as an essential process to enhance proteome diversity.