Targeted Cks1 overexpression is not sufficient to induce B cell lymphoproliferation or lymphoma.

<p>Data from two experiments (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037433#pone-0037433-g002" target="_blank">Figure 2</a>) are combined. One experiment with 2×10⁶ transplanted cells and an infection rate of 15% (n = 4 or 5 recipient mice) and one with 1×10⁶ cells and an infection rate of 45% (n = 2 or 3 recipient mice). <b>A</b>, Flow cytometric analysis of the percentage of GFP-positive B cells. Leucocytes were gated on B220+ cells. The analysis shows a divergence in the groups, some animals loose GFP positivity in the B cell compartment over time irrespective of the transduction efficiency. <b>B</b>, Left panel: Flow cytometric results of the bone marrow (BM) and spleen (S) for GFP positivity (of all cells, after lysis of erythrocytes). Right panel: percentage of GFP-positive cells in the B220 compartment of bone marrow (BM) and spleen (S). The analysis was performed on the day of indisposition and sacrifice. <b>C</b>, Representative images of flow cytometric analysis of GFP-positive B cells in the bone marrow. The genetic group is indicated. <b>D</b>, Survival of the <i>CD19-Cre;Stop-Cks1-GFP</i> group (median: 444 days) was identical to the survival of the <i>CD19-Cre;Stop-GFP</i> group (median: 391 days). All mice died from irradiation induced reasons but not B cell lymphoma.</p

Ubiquitous Cks1 overexpression in the bone marrow is not sufficient to induce myeloproliferative disease or leukemia.

<p>4×10⁶ GFP-positive cells were transplanted with an infection rate of 30%. <b>A</b>, Cks1 mRNA levels in the indicated bone marrow subsets of transplanted mice were analyzed by realtime PCR at day 160 after transplantation. Shown is the relative expression of Cks1 as compared to the B cell expression of the GFP mice as a control sample set at 1. <i>GFP</i> group: n = 2; <i>Cks1-GFP</i> group: n = 3. The bars represent the mean relative expression ± standard deviation. realtime PCR was performed in duplicates. <b>B</b>, Immunoblot analysis of the bone marrow of transplanted mice on the day of indisposition and sacrifice. <b>C</b>, The left panel shows the GFP positivity of the bone marrow (BM) and spleen (S). The three right panels show the percentages of B-, T- and myeloid cells. <b>D</b>, Representative flow cytometric dot blot images of the myeloid cell compartment of <i>GFP</i>, <i>Cks1-GFP</i> and <i>Myc-GFP</i> animals on the day of their sacrifice. <b>E</b>, Survival analysis of lethally irradiated recipient mice that were transplanted with <i>GFP</i>, <i>Cks1-GFP</i> or <i>Myc-GFP</i> infected bone marrow. None of the <i>GFP</i> or <i>Cks1-GFP</i> mice died of leukemia, while all <i>Myc-GFP</i> mice succumbed to AML. Median survival: <i>GFP</i> group, 469 days; <i>Cks1-GFP</i> group, 541.5 days; <i>Myc-GFP</i> group, 52 days. Only the survival of <i>Myc-GFP</i> group is significantly different from the other groups.</p

Cks1 regulates proliferation in oncogene-induced AML by suppressing p27^Kip1.

<p><b>A</b>, 5-FU-mobilized bone morrow from mice of the indicated genotype was infected with <i>Myc-GFP</i> virus and 2×10⁶ GFP-positive cells were transplanted with an infection rate of about 40% into lethally irradiated recipients. Immunoblot analysis of the indicated proteins was performed 3 weeks after transplantation. <b>B</b>, Flow cytometric anti-BrdU staining of GFP-positive Gr1/CD11b-positive bone marrow cells. 1 mg BrdU/g body weight was intraperitoneally injected 12 hours before bone marrow harvest. The bars represent the mean ± standard deviation of n = 3 or 5 individual mice per genotype. <b>C</b>, Survival curve of mice transplanted with <i>Myc-GFP</i>-infected bone marrow of the indicated <i>Cks1</i> and <i>p27^Kip1</i> genotype. All mice died from AML. The median survival is not significantly different between the genotypes.</p