Chronic lymphocytic leukemia (CLL) outgrowth depends on signals from the microenvironment. We have previously found that in vitro reconstitution of this microenvironment induces specific variant isoforms of the adhesion molecule CD44, which confer human CLL with high affinity to hyaluronan (HA). Here, we determined the in vivo contribution of standard CD44 and its variants to leukemic B-cell homing and proliferation in Tcl1 transgenic mice with a B-cell-specific CD44 deficiency. In these mice, leukemia onset was delayed and leukemic infiltration of spleen, liver, and lungs, but not of bone marrow was decreased. Competitive transplantation revealed that CLL homing to spleen and bone marrow required functional CD44. Notably, enrichment of CD44v6 variants particularly in spleen enhanced CLL engraftment and proliferation, along with increased HA-binding. We recapitulated CD44v6 induction in the human disease and revealed the involvement of MAP kinase and NF-kB signaling upon CD40 Ligand and B-cell receptor stimulation by in vitro inhibition experiments and chromatin immunoprecipitation assays. The investigation of downstream signaling after CD44v6-HA engagement uncovered the activation of ERK and p65. Consequently, anti-CD44v6 treatment reduced leukemic cell proliferation in vitro in human and mouse confirming the general nature of the findings. In summary, we propose a CD44-NF-kB-CD44v6 circuit in CLL, allowing tumor cells to gain HA binding capacity and supporting their proliferation.