Chronic lymphocytic leukemia (CLL) is a variable disease; therefore, markers to identify aggressive forms are essential for patient management. Here, we have shown that expression of the costimulatory molecule and microbial sensor SLAMF1 (also known as CD150) is lost in a subset of patients with an aggressive CLL that associates with a shorter time to first treatment and reduced overall survival. SLAMF1 silencing in CLL-like Mec-1 cells, which constitutively express SLAMF1, modulated pathways related to cell migration, cytoskeletal organization, and intracellular vesicle formation and recirculation. SLAMF1 deficiency associated with increased expression of CXCR4, CD38, and CD44, thereby positively affecting chemotactic responses to CXCL12. SLAMF1 ligation with an agonistic monoclonal antibody increased ROS accumulation and induced phosphorylation of p38, JNK1/2, and BCL2, thereby promoting the autophagic flux. Beclin1 dissociated from BCL2 in response to SLAMF1 ligation, resulting in formation of the autophagy macrocomplex, which contains SLAMF1, beclin1, and the enzyme VPS34. Accordingly, SLAMF1-silenced cells or SLAMF1lo primary CLL cells were resistant to autophagy-activating therapeutic agents, such as fludarabine and the BCL2 homology domain 3 mimetic ABT-737. Together, these results indicate that loss of SLAMF1 expression in CLL modulates genetic pathways that regulate chemotaxis and autophagy and that potentially affect drug responses, and suggest that these effects underlie unfavorable clinical outcome experienced by SLAMF1lo patients.