Efficient bone marrow (BM) homing is a prerequisite for successful engraftment of transplanted hematopoietic progenitor cells (HPC). Contradictory conclusions about the dependence of homing on SDF-1/CXCR4 have clouded our understanding of its participation in homing and cooperation with other defined molecular pathways. Based on the principle that homing is the coordinated outcome of several pathways, we sought to unravel cooperative and compensatory molecular mechanisms guiding BM homing of BM-HPC. BM homing of fresh BM-HPC does not require SDF-1/CXCR4 or other Gi-protein signals, since fresh BM-HPC rendered either SDF-1 unresponsive (by AMD3100, evaluated 3h after transplantation) or Gi-signaling refractory (by Pertussis toxin (PTX), evaluated 18h after transplantation) homed similarly to untreated control BM-HPC (0.56±0.03% vs. 0.60±0.01% of injected CFU-C recovered per femur (−/+AMD3100 @3h); 0.89±0.08% vs. 0.80±0.07% (−/+PTX @18h)). This was likely due to compensation by α4-integrin interacting with VCAM-1: a contribution of SDF-1/CXCR4- or Gi-protein-mediated signals to BM homing became apparent when inhibition was exerted on α4−/− HPC, significantly augmenting the intrinsic homing defect of α4−/− HPC (0.31±0.02% vs. 0.11±0.03% (−/+AMD3100 @3h); 0.52±0.08% vs. 0.02±0.01% (−/+PTX @18h) (p<0.005)). Similar conclusions were revealed when VCAM-1 deficient hosts were used as recipients of PTX-treated BM-HPC (p<0.005 compared to VCAM-1−/− recipients of untreated control BM-HPC), but not when E/P-selectin−/− recipients were used. Thus Gi-signals do not synergistically interact with E/P-selectins, although α4 integrins do so for the homing of fresh BM-HPC (p<0.005 compared with α4−/− to WT or WT to E/P−/− transplantation). When cytokine incubated BM cells were used for transplantation (SCF, 100 ng/mL for 16–18 h) the hierarchy of molecular pathway usage in homing was shifted towards a dominance of SDF-1/CXCR4 and other Gi-signals (0.60±0.05% vs. 0.37±0.08% (−/+AMD3100 @3h, p<0.05); 0.74±0.05% vs. 0.16±0.02% (−/+PTX @18h, p<0.005)), with only partial compensation through α4/VCAM-1 and endothelial selectins. Of note, expression of α4-integrin on short-term SCF-incubated HPC was not appreciably different from untreated controls. Moreover, SCF+PTX-treated α4−/− HPC homing was virtually abrogated (p<0.05 compared to similarly treated WT HPC). E/P−/− recipients supported normal homing of SCF-incubated HPC, but homing of SCF+PTX-incubated HPC in E/P-selectin−/− recipients was further decreased (0.02±0.00%, p<0.005). Additional homing studies confirmed that these findings likewise apply to SCF-incubated c-kit+lin− cells, a population enriched for stem cells, and that this functional phenotype is not specific to SCF, since it was observed for TPO±PTX incubated BM-HPC. These studies suggest a flexible hierarchy of cooperating homing pathways, in which dominant players are repositioned with changing cytokine milieu, and possibly source of HPC.