Whole transcriptome sequencing of bovine mammary epithelial subpopulations

The bovine mammary gland has a heterogeneous epithelial population which comprises terminally differentiated luminal and myoepithelial cells but also bipotent or lineage restricted progenitors and adult stem cells. The aim of our study was to use a novel surface marker (P-Cadherin) to characterize different mammary subpopulation, to sort adult stem cells with better enrichment and to perform whole RNA-seq among sorted population to identify biological processes or molecular functions associated gene enrichment. We found that only the CD49fhigh/PCadherinneg was enriched for adult mammary stem cells, while in other fractions we could detect luminal progenitors. Analysis of functional enrichments showed that in the stem cell compartment proliferation associated genes were downregulated, while genes relating to adhesion to ECM and to other neighboring cells were upregulated. Bovine mammary cells were collected at the slaughter house from 3 animals. Isolated mammary epithelial cells of each animal were sorted into 4 fraction according to their expression of CD49f and P-Cadherin. RNA was extracted from cells for each fraction and sequence using Illumina TruSeq with a depth/coverage of 30M reads per sample. Data from the 3 animals were then used as replicates to identify differentially expressed genes among the 4 mammary subpopulations.