The steroid receptor coactivator SRC3 is essential for the transcriptional activity of oestrogen receptor α. Breast cancer(BC) pathogenesis relies upon SRC3, which also has been implicated in endocrine resistance. SRC3 is posttranslationally modified by phosphorylation, but these events have not been investigated with regard to functionality or disease association. Here we investigate the spatial selectivity of SRC3-pS543/DNA binding over the human genome and its expression in primary human BC in relation with outcome. Experimental Design Chromatin Immunoprecipitation, coupled with sequencing (ChIP-seq) was used to determine the chromatin binding patterns of SRC3-pS543 in the BC cell line MCF-7 and two untreated primary breast cancers. Immunohistochemistry was used to assess the expression of SRC3 and SRC3-pS543 in 1650 primary breast cancers. The relationship between the expression of SRC3 and SRC3-pS543, disease free survival (DFS) and breast cancer specific survival (BCSS) was assessed. Results While total SRC3 is selectively found at enhancer regions, SRC3-pS543 is recruited to promoters of ERα responsive genes, both in the MCF-7 cell line and primary breast tumour specimens. SRC3-pS543 was associated with both improved DFS (p=0.003) and BCSS (p=0.001) in tamoxifen untreated high-risk patients, such a correlation was not seen in tamoxifen-treated cases, the interaction was statistically significant (p=0.001). Multivariate analysis showed SRC3-pS543 to be an independent prognostic factor. Conclusion Phosphorylation of SRC3 at S543 affects its genomic interactions on a genome-wide level, where SRC3-pS543 is selectively recruited to promoters of ERα-responsive genes. SRC3-pS543 is a prognostic marker, and a predictive marker of response to endocrine therapy.