HS1 status is associated with differential LYN phosphorylation in CLL cells. (A) Total LYN tyrosine phosphorylation (upper panels) was evaluated after IP with an anti(α)-LYN antibody and WB for total phosphotyrosines on CNTR and HS1-KD MEC1 cell lines, showing LYN phosphorylation only in CNTR cells. Total LYN protein levels (lower panels) are detected by WB, as a control. (B) Total LYN tyrosine phosphorylation (upper panels) was detected as in (A) on primary CLL samples (patient numbers are the same as in supplemental Tables 1 and 2) with different 2DE-detected HS1 phosphorylation status (HS1^hypo-p, HS1^hyper-p), showing higher LYN phosphorylation in HS1^hypo-p samples. Total LYN protein levels (lower panels) were detected by WB, as a control. (C) In the dot plot, densitometric analysis of total LYN tyrosine phosphorylation, detected as in (A), are shown from the entire cohort of 63 primary samples, divided into 2 groups based on 2DE-detected HS1 phosphorylation status (n = 36 HS1^hypo-p vs n = 27 HS1^hyper-p). The numbers refer to the representative samples displayed in (B). Means ± SEM of the OD ratio between total LYN tyrosine phosphorylation and total LYN protein levels are shown. Significantly higher total LYN phosphorylation was observed in HS1^hypo-p samples, as compared with HS1^hyper-p. **P ≤ .01, Mann-Whitney U test. P-Y, phosphotyrosines.