Abstract Peripheral monocytes, consisting of distinct Ly6C+ and Ly6Cneg subsets in mice, are innate immune effector cells that exhibit a relatively short half-life of 1–3 days. While the differentiation and function of these monocyte subpopulations have been explored extensively, the cellular mechanisms regulating the maintenance and turnover of these populations are not completely understood. The focus of our current study is to investigate the role of proline-rich tyrosine kinase 2 (Pyk2) in the establishment and/or maintenance of monocyte populations at steady-state. Using a Pyk2 knock-out mouse model (Pyk2−/−), we show for the first time that Pyk2−/− mice exhibit reduced bone marrow (BM) cellularity, resulting in a reduction of total BM monocytes in these mice. Accordingly, Pyk2−/− mice display reduced numbers of BM monocyte progenitors and Ly6C+ monocytes; however, there is no reduction in the representation of Ly6Cneg monocytes in the absence of Pyk2, suggesting that Pyk2 regulates the relative proportion of monocyte subsets normally represented in the BM at steady state. In support of this conclusion, a similar phenotype was observed in the peripheral blood and spleen. Data from reciprocal and mixed BM chimera experiments indicate that the alterations in monocyte populations exhibited by Pyk2−/− mice are due to factors intrinsic to the monocytes. Pyk2 does not appear to regulate the proliferation, differentiation, or egress of monocyte subsets. Instead, experiments using annexin V and cleaved caspase as markers for apoptosis indicate that Pyk2 may promote apoptosis in BM monocytes, thereby acting as an important regulator of turnover in these short-lived effector cells.