Transcript Profiling of Elf5+/- Mammary Glands During Pregnancy Identifies Novel Targets of Elf5

Elf5, an epithelial-specific Ets transcription factor, plays a crucial role in the pregnancy-associated development of the mammary gland. However, the molecular mechanisms employed by Elf5 to exert its effects on the mammary gland are largely unknown. Transcript profiling was used to investigate the transcriptional changes that occur as a result of Elf5 haploinsufficiency. We show that the development of the Elf5+/- gland is delayed at a transcriptional and morphological level, due to the delayed increase in Elf5 protein in these glands. We also identify a number of potential Elf5 target genes, including Mucin 4, whose expression, is directly regulated by the binding of Elf5 to an Ets-binding site within its promoter. We identify novel transcriptional targets of Elf5 and show that Muc4 is a direct target of Elf5, further elucidating the mechanisms through which Elf5 regulates proliferation and differentiation in the mammary gland. We used Compugen 22,000 oligo arrays from the Adelaide Microarray Centre to determine the transcriptional effects of the loss of one Elf5 allele on mammary gland development. We examined Elf5+/+ and Elf5+/- mammary glands over 5 timepoints of mammary gland development in three experiments. Each experiment used the pooled RNA of 2 mice, resulting in a total of 6 individual mice at each timepoint per genotype. Replica 1 of each condition was labelled with the one fluorophore, and replicas 2 and 3 with the other fluorophore. A common reference design was used for this experiment. RNA extracted from eight pooled 17.5dpc C57BL/6 mouse embryos was used as the reference sample. In total, we performed 30 micraoarray hybridisations, examining 5 timepoints of mammary gland development in 2 genotypes (Elf5+/+ and Elf5+/-). This experiment was repeated with a total of three biological replicates.