Collagen receptors GPVI and integrin α2β1 are highly expressed on blood platelets and megakaryocytes (MKs), their immediate precursors. After vessel injury, subendothelial collagen becomes exposed and induces platelet activation to prevent blood loss. Collagen types I and IV are thought to have opposite effects on platelet biogenesis, directing proplatelet formation (PPF) into the blood to prevent premature release within the marrow cavity. We used MKs lacking collagen receptors or treated MKs with blocking antibodies and could demonstrate that collagen I-mediated inhibition of PPF is specifically mediated by GPVI. Other collagen types compete with binding and diminish the inhibitory signal, which was entirely dependent on receptor-proximal Src family kinases, while Syk and LAT were dispensable. Adhesion assays indicate that MK-binding to collagens is mediated by α2β1 and that collagen IV at the vascular niche might displace collagen I from MKs and thus contribute to prevention of premature platelet release into the marrow cavity and thereby directionally promotes PPF at the vasculature.