Drosophila myeloid leukemia factor acts with DREF to activate the JNK signaling pathway

Area of Science:

• Molecular biology
• Genetics
• Developmental biology

Background:

• Drosophila myelodysplasia/myeloid leukemia factor (dMLF), a homolog of human MLF1, is an oncogene.
• Overexpression of dMLF in Drosophila wing imaginal discs induces apoptosis and an atrophied wing phenotype.
• The precise molecular mechanisms underlying dMLF-induced apoptosis are not fully understood.

Purpose of the Study:

• To elucidate the molecular mechanisms by which dMLF induces apoptosis in Drosophila.
• To investigate the role of the Jun N-terminal kinase (JNK) pathway in dMLF-mediated phenotypes.
• To identify the interaction between dMLF, DNA replication-related element-binding factor (DREF), and the basket (bsk) gene.

Main Methods:

• Yeast two-hybrid screen to identify interacting proteins.
• Genetic manipulation using engrailed-GAL4 driver and loss-of-function mutations.
• Reporter gene assays (puckered-lacZ) to monitor JNK activation.
• Chromatin immunoprecipitation (ChIP) and quantitative real-time PCR (qPCR) to assess gene binding and expression.
• Transient luciferase expression assays to measure promoter activity.

Main Results:

• Loss-of-function mutation in Drosophila JNK (basket, bsk) suppressed the dMLF-induced atrophied wing phenotype.
• dMLF overexpression induced ectopic JNK activation and apoptosis in wing discs.
• dMLF binds to the bsk promoter region containing the DREF-binding consensus sequence (DRE).
• Knockdown of dMLF reduced bsk gene promoter activity.
• dMLF interacts with DREF in vivo.

Conclusions:

• dMLF acts in concert with DREF to activate the bsk promoter.
• This activation leads to JNK pathway stimulation and subsequent apoptosis.
• The dMLF-DREF-JNK-bsk axis is a key pathway regulating apoptosis and development in Drosophila wing imaginal discs.