Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2016 Sep 27;17(1):249-260.
doi: 10.1016/j.celrep.2016.08.076.

Truncating Prolactin Receptor Mutations Promote Tumor Growth in Murine Estrogen Receptor-Alpha Mammary Carcinomas

Affiliations
Free PMC article

Truncating Prolactin Receptor Mutations Promote Tumor Growth in Murine Estrogen Receptor-Alpha Mammary Carcinomas

Obi L Griffith et al. Cell Rep. .
Free PMC article

Abstract

Estrogen receptor alpha-positive (ERα+) luminal tumors are the most frequent subtype of breast cancer. Stat1(-/-) mice develop mammary tumors that closely recapitulate the biological characteristics of this cancer subtype. To identify transforming events that contribute to tumorigenesis, we performed whole genome sequencing of Stat1(-/-) primary mammary tumors and matched normal tissues. This investigation identified somatic truncating mutations affecting the prolactin receptor (PRLR) in all tumor and no normal samples. Targeted sequencing confirmed the presence of these mutations in precancerous lesions, indicating that this is an early event in tumorigenesis. Functional evaluation of these heterozygous mutations in Stat1(-/-) mouse embryonic fibroblasts showed that co-expression of truncated and wild-type PRLR led to aberrant STAT3 and STAT5 activation downstream of the receptor, cellular transformation in vitro, and tumor formation in vivo. In conclusion, truncating mutations of PRLR promote tumor growth in a model of human ERα+ breast cancer and warrant further investigation.

Keywords: PRLR; STAT1; breast cancer; estrogen-receptor positive; luminal; mouse model; whole genome sequencing.

Figures

Figure 1
Figure 1. Sample summary
Samples from wild type 129/Sv (+/+) and Stat1 knockout (−/−) were used for whole genome sequencing in discovery and subsequent extension for targeted sequencing of Prlr. Mice with matched normal tail DNA used for analysis are indicated with a red tail. Ductal carcinoma in situ (DCIS) samples were prepared from formalin-fixed, paraffin embedded (FFPE) samples. See also Table S7 and Figure S2.
Figure 2
Figure 2. Mutational hotspot analysis of Prlr
The diagrams depict the full-length 608-amino acid residue coding region of Prlr (ENSMUST00000124470; Ensembl v67) that is encoded by 9,900 base pairs. Panel A depicts mutations identified in the original discovery set of tumors by whole genome sequencing and panel B depicts those identified in the extension/validation set (including DCIS samples) by Sanger/MiSeq sequencing. A total of 32 frame-shift deletions, 3 frame-shift insertions, 4 nonsense SNVs and 1 in-frame deletion introducing a stop codon were identified in Prlr that cluster in 2 hotspots around residues 318 and 330. See also Figure S4, Table S5, Table S10, Appendix 1, Appendix 2, Appendix 3, and Appendix 4.
Figure 3
Figure 3. Co-expression of full-length and truncated Prlr promote Stat3 and Stat5 activation, cellular transformation in vitro and tumor formation in vivo
(A) Stat1−/− MEFs expressing full-length (FL), truncated (T) Prlr or both (FL/T) were stained for phosphorylated Stat3 (pStat3) or phosphorylated Stat5 (pStat5) (y-axis). Rabbit (Rb) IgG was used as an isotype control. MEFs also expressed mJak.IRES.GFP (x-axis) to mediate signaling downstream of the Prlr proteins. (B) Stat1−/− MEFs expressing the indicated Prlr constructs were analyzed by anchorage-independent soft agar assay. The number of colonies was counted after single cells had been cultured for 3 weeks. (C) Stat1−/− MEFs alone (−), Stat1−/− MEFs transduced with vector alone (Jak2) or vector expressing full-length (FL/FL-Jak2), truncated (T/T-Jak2) or both (FL/T-Jak2) Prlr were implanted into nude mice. Stat1−/− MEFs expressing Kras were used as positive control. Tumor growth was monitored over time. The percentages of animals that developed palpable tumors in each experimental group were plotted. * p<0.002, ** p<0.0001. See also Figure S3, Figure S5.
Figure 4
Figure 4. PRLR isoform usage versus STAT1 expression in human TCGA breast cancer RNA-seq data
STAT1 expression levels from TCGA RNA-seq data were binned into tertiles (low, medium, high) (mid panels) and separated into luminal or basal/Her2 breast cancer subtypes. The ratios of full-length (FL) to truncated (T) PRLR isoform expression (FL/T ratio values) calculated in terms of junction per million (JPM) were compared using a Wilcoxon rank sum test with continuity correction between low and high STAT1 expressing groups (top panels). For reference, read counts of FL and T are plotted (bottom panels). See also Figure S6 and Table S6.

Similar articles

See all similar articles

Cited by 7 articles

See all "Cited by" articles

Publication types

MeSH terms

LinkOut - more resources

Feedback