Analysis of Gene Expression in PTHrP-/- Mammary Buds

Analysis of Gene Expression in PTHrP-/- Mammary Buds Supports a Role for BMP Signaling and MMP2 in the Initiation of Ductal Morphogenesis. Parathyroid hormone-related protein (PTHrP) acts on the mammary mesenchyme and is required for proper embryonic mammary development. In order to understand PTHrP's effects on mesenchymal cells, we profiled gene expression in WT and PTHrP-/- mammary buds, and in WT and K14-PTHrP ventral skin, at E15.5. By cross-referencing the differences in gene expression between these groups, we identified 35 genes potentially regulated by PTHrP in the mammary mesenchyme, including 6 genes known to be involved in BMP signaling. One of these genes was MMP2. We demonstrated that PTHrP and BMP4 regulate MMP2 gene expression and MMP2 activity in mesenchymal cells. Using mammary bud cultures, we demonstrated that MMP2 acts downstream of PTHrP to stimulate ductal outgrowth. Future studies on the functional role of other genes on this list should expand our knowledge of how PTHrP signaling triggers the onset of ductal outgrowth from the embryonic mammary buds. Five experimental groups (WT mammary buds, PTHrP-/- mammary buds, WT ventral skin, K14WT ventral skin, K14-PTHrP ventral skin). Three replicates in each group. Differentially expressed genes are listed in the following supplementary files: GSE17654_List1a-Higher_in_KO_Buds_than_WT_Buds.txt GSE17654_List1b-Lower_in_KO_Buds_than_WT_Buds.txt GSE17654_List2a-Higher_in_WT_Skin_than_WT_Buds.txt GSE17654_List2b-Lower_in_WT_Skin_than_WT_Buds.txt《PTHrP敲除（PTHrP-/-）乳腺芽的基因表达分析证实骨形态发生蛋白（BMP）信号通路与基质金属蛋白酶2（MMP2）在导管形态发生启动中发挥作用》。甲状旁腺激素相关蛋白（PTHrP）可作用于乳腺间充质，是胚胎乳腺正常发育所必需的调控因子。为解析PTHrP对间充质细胞的调控效应，本研究在胚胎发育第15.5天（E15.5）对野生型（WT）与PTHrP敲除（PTHrP-/-）乳腺芽，以及野生型（WT）与角蛋白14启动子驱动的PTHrP过表达（K14-PTHrP）腹侧皮肤样本开展了基因表达谱分析。通过比对各组间的基因表达差异，我们筛选出乳腺间充质中35个可能受PTHrP调控的基因，其中包含6个已被证实参与BMP信号通路的基因，MMP2即为其中之一。本研究证实，PTHrP与BMP4可在间充质细胞中调控MMP2的基因表达与酶活性。通过乳腺芽体外培养实验，我们证实MMP2可作为PTHrP的下游效应因子，促进导管向外生长。后续针对该列表中其余基因功能的研究，将进一步完善我们对PTHrP信号通路如何触发胚胎乳腺芽导管向外生长起始过程的认知。本研究共设置5组实验样本：野生型乳腺芽、PTHrP敲除乳腺芽、野生型腹侧皮肤、K14野生型腹侧皮肤以及K14-PTHrP腹侧皮肤，每组设置3次生物学重复。差异表达基因详见以下补充文件：GSE17654_List1a-Higher_in_KO_Buds_than_WT_Buds.txt、GSE17654_List1b-Lower_in_KO_Buds_than_WT_Buds.txt、GSE17654_List2a-Higher_in_WT_Skin_than_WT_Buds.txt、GSE17654_List2b-Lower_in_WT_Skin_than_WT_Buds.txt。