Background: Among patients with estrogen receptor (ER)-positive breast cancer, progesterone receptor (PR)-negative tumors were shown to have worse prognosis than PR-positive tumors. However, PR-negative tumors were underrepresented in trials such as TAILORx and RxPONDER, and the role of PR status in the setting of 21-gene recurrence score (RS) remains unclear. We performed an observational cohort study to evaluate the association of PR status with RS and the magnitude of chemotherapy benefits on survival. Methods: The National Cancer Database (NCDB) was queried for women diagnosed between 2010 and 2017 with ER-positive, human epidermal growth factor receptor 2 (HER2)-negative, pT1-3N0-1a breast cancer who underwent surgery and endocrine therapy. Logistic and Cox multivariable analyses (MVA) were used to identify variables associated with high RS (> 25) and overall survival (OS), respectively. Interaction test was performed among PR status, chemotherapy, and nodal staging. Propensity score matching was performed to reduce selection bias. Sensitivity analysis was performed after excluding those with postdiagnosis survival of less than 6 months to reduce immortal time bias. Results: A total of 143,828 patients met our criteria (n = 110,421 for PR-positive/pN0, n = 11,897 for PR-negative/pN0, n = 19,928 for PR-positive/pN1a, n = 1,582 for PR-negative/pN1a). Median follow up was 51.5 months (interquartile range 34.8-71.9). On logistic MVA, PR-negative tumors were more likely to have high RS (adjusted odds ratio [aOR] 6.68, 95% confidence interval [CI] 6.40-6.97, p < 0.001). On Cox MVA, PR-negative tumors were associated with worse OS (adjusted hazards ratio [aHR] 1.20, 95% CI 1.10-1.31, p < 0.001). Interaction among PR status, chemotherapy, and nodal staging was statistically significant (interaction p = 0.049). On subgroup analyses, the magnitude of chemotherapy benefit for OS was comparable among pN0 tumors (PR-positive: aHR 0.74, 95% CI 0.66-0.82; PR-negative: aHR 0.63, 95% CI 0.51-0.77) and was greater for PR-negative status among pN1a tumors (PR-positive: aHR 0.57, 95% CI 0.47-0.67; PR-negative: aHR 0.31, 95% CI 0.20-0.47). Similar findings were noted in 9,979, 1,822, 4,196, and 354 matched pairs for PR-positive/pN0 (HR 0.43, 95% CI 0.37-0.50), PR-negative/pN0 (HR 0.53, 95% CI 0.41-0.69), PR-positive/pN1a (HR 0.55, 95% CI 0.45-0.67), and PR-negative/pN1a (HR 0.25, 95% CI 0.14-0.45) tumors, respectively. On sensitivity analysis, our findings were consistent in Cox MVA using interaction and subgroup analyses. Conclusions: To our knowledge, this is the largest study using a nationwide oncology database suggesting that PR-negative status is an independent, adverse prognostic factor for survival associated with high RS, with greater chemotherapy benefits compared to PR-positive status among pN1a tumors even after adjusting for RS.