Entry of PTEN-Long into 786-0 cells.

<p><b>A</b>, 786-0 cells treated with 100 nM purified PTEN-Long-GFP or PTEN-GFP for 5 h and imaged, showing PTEN-Long-GFP but not PTEN-GFP entered 786-0 cells. The scale bar represents 10 µm. <b>B</b>, Immunoprecipitation for PTEN on cell lysates pretreated with PTEN-Long or PTEN followed by Western blotting analysis with antibody against PTEN, showing only PTEN-Long was detected.</p

Overexpression of PTEN-Long inhibits cell migration and reduces cell invasion.

<p><b>A</b>, 786-0 cells stably expressing indicated proteins were subjected to <i>in vitro</i> wounding and followed by phase-contrast imaging. The fields in between the white lines indicate the wound track. The scale bar represents 20 µm. <b>B</b>, The number of cells migrated in the wound track from 6 random separate fields were counted for each cell and expressed as a mean ± SD (error bars). <b>C</b>, Invasive properties of 786-0 cells were inhibited by 30% by PTEN overexpression and by 25% by PTEN-Long overexpression, as compared to empty vector control cells.</p

Protein expression of PTEN-Long was significantly reduced or lost in ccRCC.

<p><b>A</b>, Western blot analysis of PTEN, PTEN-Long, and pAkt. Represented are 6 pairs of clear cell renal cell carcinomas and their corresponding normal renal tissues. In clear cell renal cell carcinomas the PTEN-Long expression was either negative or very weak, compared to normal renal tissue. N, normal renal tissue; T, tumor tissue. <b>B</b>, The ratio of PTEN, PTEN-Long, Akt, and pAkt to Actin in 45 samples. The PTEN-Long expression in ccRCC was reduced significantly in comparison with the corresponding normal renal tissue. Note that tumor samples with undetectable PTEN-Long and PTEN were excluded from quantification.</p

PTEN-Long treatment in culture inhibits PI3K signal and induces apoptosis in 786-0 cells.

<p><b>A</b>, Coomassie stained gel showing representative protein preparations of PTEN, PTEN^G129R, PTEN-Long, and PTEN-Long^G302R. <b>B</b>, Effect of treatment with 25 nM PTEN-Long or other related proteins for 1h on pAkt and pPRAS40. PTEN-Long but not other proteins reduced intracellular phosphorylation of the Akt and PRAS40 in 786-0 cells. <b>C</b> and <b>D</b>, Effect of treatment with PTEN-Long or related proteins at different doses for 24 h on serum-starved 786-0 cells showed that PTEN-Long but not PTEN suppressed PI3K signaling and induced apoptosis, indicated by cleavage of caspase 3 in a dose-dependent manner. Percentages of apoptotic cells were also determined by PI/FACS (<b>C</b>). n = 4; bars represent the means ± SD. <b>E</b>, Representative pictures of the treated cells were taken by phase contrast photography. Protein concentrations were 50 nM. The scale bar represents 20 µm.</p

Overexpression of PTEN-Long inhibits cell proliferation and induces cell death.

<p><b>A</b>, Cell proliferation assay indicating reduced growth in 786-0 cells expressing PTEN or PTEN-Long compared to PTEN^G129R, PTEN-Long^G302R, or empty vector transfected cells. <b>B</b>, AnnexinV staining followed by flow cytometry analysis indicating overexpression of PTEN or PTEN-Long promoted serum starvation-induced apoptosis.</p

PTEN-Long inhibits the proliferation of 786-0 cells in xenograft mouse model.

<p><b>A</b>, 786-0 cells engrafted nude mice were treated with 5 mg/kg of PTEN or PTEN-Long for 6 days. PTEN-Long but not PTEN inhibited tumor growth. <b>B</b>, Graph of tumor diameters as measured by calipers and treated with either PTEN or PTEN-Long at 5 mg/kg for different time periods. n = 6; bars represent the means ± SD.</p

Overexpression of PTEN-Long suppressed PI3K-Akt signaling in 786-0 cells.

<p>Western blot analysis of lysates from 786-0 cells showing overexpression of PTEN or PTEN-Long, but not PTEN^G129R or PTEN-Long^G302R, suppressed Akt phosphorylation (Ser473) and its downstream event.</p

Additional file 2 of GhNFYA16 was functionally observed positively responding to salt stress by genome-wide identification of NFYA gene family in cotton

Additional File 2: Table S3 Cis-acting elements of upland cotton NFYA family members

Additional file 1 of GhNFYA16 was functionally observed positively responding to salt stress by genome-wide identification of NFYA gene family in cotton

Additional File 1: Figure S1 Distribution of NFYA family members in 11 species. Figure S2 Analysis of Non-synonymous (Ka) to Synonymous (Ks) ratio. Table S1 NFYA family genes correspond to eleven species gene renames. Table S2 Analysis of the physical and chemical properties of NFYA genes in G.hirsutum

Additional file 3 of GhNFYA16 was functionally observed positively responding to salt stress by genome-wide identification of NFYA gene family in cotton

Additional File 3: Table S4 Expression values of 12 selected genes under salt stress