An total maximize in b catenin protein ranges with a resul tant transfer of b catenin on the nucleus was observed in cells treated with or over expressing OPN. Via the nuclear import of b catenin, OPN increases both the transcription and protein ranges of MMP 7 and CD44, that are recognized TCF LEF transcription targets, The Erk pathway is among the best studied selleck chemicals MG-132 MAPK pathways in mammals and continues to be shown to be deregulated in roughly a single third of all human cancers, Erk1 2 activation regulates proliferation, differentiation, survival, migration, angiogenesis, as well as chromatin remodeling as a result of the phosphorylation of the two cytoplasmic and nuclear targets such as phos phatases, transcriptional variables, and cytoskeletal professional teins, Within the canonical Erk1 two pathway, receptor tyrosine kinases are activated by specific ligands and set off guanosine trisphosphate loading of your Ras protein, which may then recruit the Raf kinases, These kinases consecutively phos phorylates and activates MEK, ulti mately leading to the activation of Erk1 2.
In addition to this pathway, Erk1 2 continues to be proven to be activated by a range of pathways determined by the personal ligand, cell surface receptor, and cell style, Das et al. previously demonstrated that OPN induces AP 1 activa tion and uPA secretion via c Src EGFR Erk signal ing in breast cancer cells which eventually handle the motility in these cells, As a result of existence of wide variation in the pathways resulting in selleck chemicals Erk1 two activation, we investigated the OPN induced signaling pathway which lead to Erk1 two activation in prostate cancer cells as well as purpose of cell surface receptors on this procedure. Raf is targeted for the plasma membrane upon activa tion by a tiny GTPase.
Phosphorylation of c Raf at ser ine 259 is definitely an inhibitory event occurring via Akt, Former studies have shown that osteoclast survi val is dependent within the Erk1 2 signaling pathway, Improved osteoclast production and exercise contributes to excessive bone reduction in ailments such as osteoporosis and tumor induced osteolysis, which continues to be linked to prostate cancer, Mainly because prostate cancer outcomes in metastases for the bone in about 80% of autopsied scenarios, prostate cancer cells present a logical technique by which to study the relationships of bone extracellular matrix proteins and tumorigenesis, OPN acts like a paracrine and autocrine mediator of prostate cancer development and progression, OPN purpose inside the activation of MAPK pathway needs more elucidation.

Erk1 2 phosphorylation of MiTF played a significant part in activating p21WAF1 CIP1 transcription along with a temporary G1 cell cycle arrest, which enhanced cell survival just after UVC radiation. These final results suggest a novel function of MiTF in linking Erk1 2 acti vation and p21WAF1 CIP1 regulation right after UVC radiation in normal human melanocytes and melanoma cells. Results MiTF is phosphorylated and transiently degraded just after UVC in NHMs and a few melanoma cells To examine whether or not MiTF plays a part in DNA injury response, two normal human melanocyte cell lines have been exposed to potent DNA damaging agent UVC and allowed them to recover for var ious intervals of time. As proven in Fig 1A, MiTF at base line was detected as being a doublet band on western blot. the lower band represented unphosphorylated and the major band the phosphorylated type of MiTF, 1 hour immediately after UVC, all the MiTF was shifted on the top rated band, The phosphorylation continued for two hrs just after UVC, followed by a decrease of MiTF protein at four and six hrs.
After that, MiTF protein begun to recover 9 hours submit radiation and nearly fully recovered to its pre treatment method amounts twelve to 24 hours right after UVC, The two NHMs had been isolated from neonatal foreskin of a Caucasian and an African black baby respectively. There was no significant difference inside their response to UVC. A similar response was observed in c83 2C melanoma cells, MiTF degradation was even further confirmed by immunofluorescence, purchase CC-292 c83 2C cells were exposed to UVC and fixed for immuno fluorescence staining at different time points. Consistent with its nuclear localization, the fluorescence signal for MiTF was largely observed in nuclei, However, no exact foci had been observed, nor was there a dramatic re localization from the protein at 1 hour submit radiation, suggesting that phosphorylation of MiTF was not a sig nal for recruiting DNA repair proteins to DNA damage web-sites, nor was it a signal for translocation to cytoplasm.
MiTF phosphorylation was examined one hour following var ious doses of UVC radiation. as minimal as 1 mJ cm2 of radiation led to MiTF phosphorylation in c83 2C cells, MiTF phosphorylation is by means of Erk1 2 mitogen activated protein kinases and it is demanded for its subsequent proteasome dependent degradation To investigate the upstream signal for MiTF phosphory order ONX-0914 lation, three kinase inhibitors were incubated with NHMs prior to they were exposed to UVC. MEK inhibitor U0126 which leads to Erk1 two inhibition, the p38 MAPK inhibitor SB203580, and wortamannin, an inhibitor of phosphatidy linositol 3 kinase, Ataxia telangiectasia mutated and ATM and Rad3 connected kinase. Cells were exposed to UVC and collected one hour later on to examine MiTF phosphorylation. As shown in Fig 2A, best panel, between these kinase inhibitors, only U0126 inhibited UVC mediated MiTF phosphorylation, sug gesting that Erk1 two could be the upstream kinase.

This suggests that therapies targeted to KLF5 could have potential therapeutic advantage to patients with colorectal cancer. Without a doubt, a current screen for tiny molecule inhi bitors of KLF5 expression has yielded numerous potent compounds that inhibit proliferation of colorectal cancer cells, Additional investigation may possibly demonstrate KLF5 an appealing target for intervention within the prevention or therapy of colorectal cancer. Conclusions Reduction of tumor suppressor genes and activation of onco genes are hallmarks of cancers. During the situation of colorectal cancer, loss of APC and activation of KRAS are com mon. Here, we present a robust mouse model of intest inal tumorigenesis with the generation of ApcMin KRASV12 mice. These mice show an enhanced selleck propen sity for building intestinal tumors at an early age compared to ApcMin mice.
Moreover, we were able to significantly greatly reduce tumor burden and dimension in the com pound ApcMin KRASV12 mice by lowering expression of Klf5 with genetic usually means. ApcMin KRASV12 Klf5 mice show decreased levels of Klf5 protein as well CAY10505 as b cate nin, cyclin D1 and Ki67, all regarded markers of prolifera tion and transformation. We conclude that Klf5 can be a crucial mediator of initiation and progression of intest inal tumors resulted from ApcMin and KRASV12 mutations. Procedures Reagents Antibodies utilized in the experiments were previously described, Antibodies towards KLF5 had been gener ated against a synthetic KLF5 peptide in rabbits, Anti KLF5 antibody was used at a dilution of 1.15,000 for immunohistochemistry and at 1.4,000 for Western blot analysis. Mouse mono clonal antibody towards complete b catenin was bought from Invitrogen and applied at a dilution of one.1,000 for Western blot analyses. For immunohisto chemical examination, total b catenin antibodies obtained from BD Biosciences were employed at 1.
250 dilutions. Rabbit monoclonal cyclin D1 antibodies were obtained from Biocare Healthcare and employed at one.200 dilutions in immunohistochemical analyses and 1.two,500 dilutions for Western blot examination. Anti Ki67 antibodies had been obtained from Novocastra and utilized at 1.500 dilu tions. Anti Phospho MEK1 and anti Phospho ERK1 two antibodies, applied at one.100 dilutions, have been bought from Cell Signaling Technological innovation, Mice All research involving mice are already accredited by the Emory University Institutional Animal Care and Use Committee, C57BL 6J mice heterozygous for KRASV12 expressed from a mouse villin promoter have been previously produced, Mice double heterozygous for ApcMin and Klf5 alleles had been produced as previously described, Founder C57BL 6J mice that had been hetero zygous ApcMin alleles had been mated with those that were heterozygous for Klf5 alleles, The resulting progeny generated double heterozygous Apc Min Klf5 mice.

38 genes had been up regulated in UKF NB 3rDOX20 cells and 27 genes had been down regulated in UKF NB 3rDOX20 rela tive to UKF NB three cells, The rela tively high variety of substantially differentially regulated genes in comparison to the comparisons of UKF NB 3 vers. UKF NB 3rVCR10 or UKF NB 3rCDDP1000 cells probably final results from the distinctive statistical procedures used to analyse HGU133 Plus two. 0 or AB1700 data. To even further investigate the influence of chemoresistance acquisition around the pro angiogenic possible of cancer cells, a panel of chemsensitive and chemoresistant neu roblastoma cell lines was systematically investigated for their angiogenic phenotypes. Influence of neuroblastoma cell line supernatants on endothelial cell development and survival Neuroblastoma cell lines were grown for 7 days. Then medium was eliminated, cells had been washed and protein totally free medium was additional.
Right after 48 h incubation, supernatants were collected, adjusted for the very same protein material, selleck chemical mixed inside a 1.one ratio with fresh IMDM, and FCS was additional, HUVECs have been trypsinised and suspended in the mixtures of supernatants, fresh IMDM and FCS, 103 cells suspended in 100l of respective medium had been seeded per well in 96 nicely plates. After five days, HUVEC development was examined by viability assay, HUVECs suspended in IMDM plus 10% FCS did not develop, HUVECs suspended in IMDM plus 15% FCS, 5% pooled human serum, and fundamental fibroblast development element two. 5 ng ml formed critical, closely grown monolayers, Cell viabilities have been calculated relative to favourable control. Supernatants from cell lines adapted to cytotoxic drugs induced stronger HUVEC development than supernatants from parental chemosensitive cells, Also, the neuroblastoma cell lines UKF NB 4 and Be C that were isolated as chemoresistant cell lines from patients elements induced stronger HUVEC growth compared to the chemosensitive parental cell lines UKF NB 3, UKF NB 2, or IMR 32.
Subsequently, development kinetics of HUVECs incubated with supernatants of UKF NB 3, UKF NB 3rVCR10, UKF NB 3rCDDP1000, or UKF NB 3rDOX20 cells had been in contrast confirming enhanced development of HUVECs incubated with supernatants of chemoresistant cells, Up coming, the influence of neuroblastoma cell culture superna tants was examined on HUVEC survival. Confluent HUVEC monolayers selleck inhibitor were washed and incubated for 48 h with supernatants of UKF NB 3, UKF NB 3rVCR10, UKF NB 3rCDDP1000, or UKF NB 3rDOX20 cells and HUVEC viability was determined. Outcomes exposed improved HUVEC viability in cultures incubated with supernatants of chemoresistant cells, Lack of development components or nutrients induces apoptosis in endothelial cells, For that reason, we investigated caspase 3 7 activation as indi cator of apoptosis in confluent HUVEC monolayers incu bated for 48 h with supernatants of UKF NB 3, UKF NB 3rVCR10, UKF NB 3rCDDP1000 cells or UKF NB 3rDOX20 cells.

Furthermore, pERK immunoreactivity was predomi nantly nuclear whereas cytoplasmic positivity was identified within a subset of tumors, This staining pattern is in line with former investigations in colon also as other neoplasms, namely, endometrial, head and neck tumors and melanomas, Moreover, total nuclear or cytoplasmic ERK staining was independent of nuclear or cytoplasmic pERK status, in our cohort.
This observation is in agreement with preceding scientific studies in non compact cell lung and endometrial cancer too as in melanomas and could been explained by the hypothesis that pERK immunopositivity could arise because of ERK hyperactivation instead of overexpression, Additionally we display that read this article large nuclear, but not cyto plasmic, ERK immunopositivity is correlated with tumor stage, Interestingly, our final results show that nuclear pERK expression parallels tumor grade and tumor stage in preserving with findings of previous research in NSCLC and prostate cancer, This observation is in contrast with a former investigation through which pERK expression was not correlated with tumor stage, whereas there are other research in a variety of other tumors that have failed to sub stantiate a beneficial correlation concerning pERK expression and classical clinicopathological parameters, Taking into consideration that in our research ordinary colorectal mucosa displays minimum nuclear pERK expression, our benefits talk in favor of the possible role of this molecule in tumor evolution at the same time as, from the acquisition of the additional aggressive pheno style in colorectal carcinogenesis. Along this line, it has been recommended that constitutively energetic ERKs are cap able of affecting gene expression, being able to influence several with the hallmarks of carcinogenesis, It may very well be hypothesised that as colorectal cancer progresses to a extra innovative disease, an increase within the activation of the MAP kinase signal transduction pathway happens, Sad to say, the statistical power on the existing investigation is reduced as a result of modest cohort of pERK favourable instances.
Additional scientific studies of larger cohorts are warranted to confirm our effects. Along this line, pERK expression has been proven to become connected with poor prognosis in colorectal carcinomas, It’s been speculated the activation with the MAPK path way initiates cellular processes, which could result KW-2449 in both favorable or worse clinical outcome, a truth due to the complex signal transduction network ERKs are involved in. In reality, adverse results is usually observed, depending on the intensity and duration of the promito genic signal, The aforementioned findings are fairly major in see from the fact that protein kinases signify a group of molecular targets characterized by a cancer specific possible, enabling the improvement of new generation chemotherapeutic agents acting as kinase inhibitors.

Microarrays and gene expression examination Using the microarrays data set normalized from our an terior review, the RMA values in the 45000 probsets have been utilised to identify differentially expressed genes in T CD8 leukemias. Genes have been picked according the fol lowing criteria. the expression signal intensity didn’t fluctuate in B leukemias versus control B cells as well as the expression signal intensity was both significantly increased, or reduced in T CD8 leukemias versus control cells, The microarray dataset was deposited at Gene Expression Omnibus beneath the accession amount GSE12581, Semi quantitative RT PCR Total RNA was reverse transcribed working with the Omniscript enzyme and the oligo pri mer.
The semi quantitative PCR reactions have been carried out using the Taq polymerase kit applying an RT reaction corresponding to ten ng of RNA samples and to 2 ng for actin, Annealing temperature selleck chemical and variety of cycles were optimized for every gene. Plasmid constructions The cDNA in the total coding area of mParm one and hParm 1 had been produced by common PCR amp lification process applying primers containing certain restriction web sites. The PCR products had been then inserted in frame inside the pEGFP N1 or pcDNA3. one Myc His A vectors. Deletions had been generated making use of certain primers that amplify the particular region of curiosity as well as PCR products inserted in frame in pEGFP N1. Cell culture NIH 3T3 and Jurkat T cells have been obtained from ATCC, NIH 3T3 cells have been grown in DMEM medium supplemented with 10% CS and Jurkat cells have been cultured in RPMI supplemented with 10% FCS, 50 U penicillin and of streptomycin were additional.
Confocal microscopy For transient transfection, Jurkat cells had been transfected with 15 ug plasmids by electroporation with the Gene Pulser Process, NIH 3T3 cells buy b-AP15 were transfected applying the polyfect reagent, Each pEGFP N1 and GFP tagged mParm 1 or hParm 1 genes had been utilized. Localization of mPARM 1 and hPARM one was performed by confocal microscopy 48 h after transfection. For cell sur face membrane co localization Jurkat cells have been pelleted 48 h just after transfection, washed in PBS and overlaid for 30 min at 37 C on polylysine coated glass slides, For co localization experiments, NIH 3T3 cells had been plated on glass coverslips, grown at 50% confluency, and transfected as described over. Right after 48 h of transfection, cells have been fixed with 4% paraformaldehyde, followed by PBS washes and permeabilization with 0. 1% Triton X one hundred in PBS. Cells have been blocked in PBS with 10% goat serum, 10% BSA and 0. 1% triton, and incubated with primary antibodies. Cover slips had been incubated with Alexa Fluor 568 conjugated sec ondary antibody, washed with PBS, mounted onto slides using Prolong Gold antifade reagent and observed by confocal microscopy.

Usual epithelial cells were grown in SFM medium underneath same conditions as the carcinoma cells. which was labelled by chemoluminescence, Twenty microgram total RNA from tumor cells and standard epithe lial cells have been separated on 1% agarose gel. After transfer of the RNA onto nylon membrane both hybridization and detection procedures have been carried out in accordance to the producers guidelines. Isolation of PTX PTX was isolated chromatographically through the marine Cnidaria Palythoa caribaeorum and purified as we described earlier, Purified PTX was lyophilized and stored at 20 C. Cytotoxicity assay Quantification of cell death and cell lysis was depending on the measurement of LDH activity launched through the cytosol of broken cells to the supernatant applying a non radioactive LDH detection kit, Cells grown to a monolayer had been incubated for 24 h from the presence or absence of PTX.
Just after centrifugation at 250xg for 10 min. the cell absolutely free culture supernatants have been collected from PTX taken care of and untreated cells and incubated description in accordance to your makers instruction. To calculate % cytoto xicity suitable controls were measured in just about every experiment. Absorbance was measured at 492 nm and 620 nm working with an ELISA reader, Clonogenic assay At day 0, HNSCC cells and normal epithelial cells have been plated in duplicate in six very well plates. One week later on, just after cells had reached confluency, they had been incubated for 24h at a variety of PTX concentrations, Subsequently, cells were washed with PBS, fixed in ethanol and stained with crystal violet, Stained cells were measured by microscopic counting randomly deciding on at least ten middle power magnification fields.
Suggest values and conventional deviation were calculated. JNK3 inhibitory assay Pyrazolourea, LY2940680 a selective inhibitor of JNK3 was obtained from Merck Calbiochem, Germany. Ordinary epithelial cells have been seeded in six well plates and cultured until confluent. The cells had been incubated with pyrazolourea at concentrations ranging from 20 nM to one hundred nM for 3 hours to inhibit the JNK3 protein kinase. Subsequently, cells had been exposed to six ng ml PTX for 24 hrs. Lastly, cell survival was determined employing the crystal violet assay. Animal experiments SCID bg bg mice were obtained from Charles River aged ten to 12 weeks, To the carcino genicity experiments a group of tumor free mice was treated by subcutaneous injection of 0. 5ng PTX within a volume of twenty ul PBS day for five days. Subsequently,the animals were observed over a period of 8 months.

Down regulation of serpinE2 expression in human colorectal cancer cells inhibits soft agarose colony formation, migration and tumor development in nude mice We up coming investigated the impact of serpinE2 knockdown on anchorage independent growth and cell migration following downregulation of serpinE2 gene expression by RNA interference in HCT116 and LoVo cells. As shown in Figure 4A, serpinE2 mRNA were considerably decreased by respectively 37% and 88% in LoVo cells expressing shSerpinE2 or shSerpinE2 and by 77% and 92% in HCT116 expressing shSerpinE2 or shSer pinE2, conversely, expression with the control shRNA had no effect on endogenous serpinE2 expres sion, Once again, the proliferation price of these cell populations was assessed when cultured on plastic. No difference was observed from the proliferation price of subconfluent cells when serpinE2 expression was downregulated, We then verified irrespective of whether the reduction in serpinE2 expression alters the skill of colon cancer cells to type colonies in soft agarose.
As shown in Figure 4C, expression of each shRNA against SerpinE2 decreased the capability of HCT116 and LoVo cells to type colonies in soft agarose. Of note, shSerpinE2 which was significantly less effective than the shRNA to reduce serpinE2 gene expression was also less productive to cut back colony formation. selleck chemicals This indicates that serpinE2 controls anchorage independent development of human colon carcinoma cells. Additionally, as observed in caMEK expressing IECs, the dimension of foci formed at post confluency was substantially decreased in serpinE2 depleted LoVo cells, The tumorigenicity of colorectal cell lines was following assessed right after subcutaneous injection in to the flank of nude mice. As shown in Figure 5A and 5B, HCT116 and LoVo cell lines induced palpable tumors by using a brief latency period of respectively 15 and ten days just after their injection.
More importantly, downregulation of serpinE2 expression with R547 741713-40-6 shSerpinE2 in these cell lines severely impaired their capability to grow as tumors in nude mice. Ultimately, in vitro transwell migration assays had been per formed to confirm the significance of serpinE2 in colon carcinoma cell migration. As illustrated in Figure 6A, serpinE2 deficiency significantly lowered HCT116 and LoVo cell migration towards the undersurface with the membrane coated or not with fibronectin or vitro nectin, The net effect of serpinE2 knockdown was also established on invasion through the use of BD Biocoat Matrigel invasion chambers, in presence of hydroxyurea. As shown in Figure 6B, the capacity of LoVo cells to invade Matrigel was also altered by ser pinE2 silencing To test the hypothesis that this altered migration and invasion capability could resulfrom a defect in cell adhe sion, adhesion power on the substrate was examined for handle and shSerpinE2 expressing LoVo cells. t

As numerous examined shRNA con structs didn’t effectively knock down the gene, we employed commercial siRNA for this cell line, which lowered MMP13 transcript amounts to approx. 33%, Western blot evaluation also confirmed a reduction while in the pro and active varieties on the protein, with 60 and 48 kDa, respectively, As an alternative to the previously performed long-term proliferation assays, we carried out a BrdU incorporation assay as being a measure of DNA replication 72 h after transfection from the respective siRNA, Knockdown of MMP13 decreased BrdU incorporation to 60%. We also observed an greater fraction of siMMP13 transfected cells inside the G0 G1 phase of the cell cycle when compared to regulate cells, Even so, the impact was weaker compared to the impact seen in presence with the MMP 9 13 inhibitor displayed in figures 3C and 5C. Potentially, this can be as a result of incomplete MMP13 knock down.
It truly is also probably the arrest is extra enhanced in starved cells which might be confronted with development stimulus and MMP inhibitor in the same time. If MMP13 is knocked down while in the normal expanding cell culture, it might block cell cycle progression in general, irrespective on the cell cycle phase. This kind of behaviour selleck Veliparib is remi niscent from the result of development element withdrawal, which may block LY2109761 the cell cycle in G1 and G2, and could level to the likelihood that MMP13 releases an unidentified growth factor. To investigate whether the possible MMP13 dependent growth marketing factor is secreted, we treated siMMP13 transfected A375 cells with condi tioned supernatant from manage siRNA transfected cells. This could considerably restore BrdU incorporation to 80% of your control, indicating the presence of the soluble development advertising aspect. In summary, these data indicate that MMP13 plays a vital purpose from the development component induced prolifera tion of melanocytes and melanoma cells at the same time as inside the dedifferentiation of melanocytes. Discussion In many melanomas, MMPs are aberrantly expressed. All MMPs upregulated in Hm cells were previously reported to become made in melanoma, specifically MMP1 and 9, The bring about of MMP expression in melanoma is largely unknown, but constant ERK sig nalling, e.

The propensity for the ApcMin, Apc 1638, KRASV12, Apc 1638 KRASV12 mice to create tumors inside the smaller intestine rather then the colon has become reported, It can be of interest to note that there’s a distinction in regional dis tribution of small bowel tumors involving ApcMin and ApcMin KRASV12 mice tumors from the former mice were additional distally distributed when those within the latter have been extra proximally distributed, This big difference in tumor distribution will not seem to become as a result of regional variations in expression of your KRASV12 transgene from your villin promoter, The result of KRASV12 allele introduction about the shift in tumor distribution additional proximally is consequently not clear at this time.
A related trend towards distribution of smaller bowel tumors in the ApcMin mice has been reported, We just lately reported the significant part for Klf5 in tumor initiation in ApcMin mice, Klf5 haploinsufficiency in ApcMin mice resulted inside a signifi cant decrease in tumor quantity and dimension, Results selleck chemicals BAY 11-7082 in the current research show a related effect on tumor formation at 12 weeks of age in ApcMin KRASV12 mice that have been heterozygous for your Klf5 alleles, using the intestinal tumor burden lowered by greater than 90% in the triple ApcMin KRASV12 Klf5 transgenic mice when compared to the double ApcMin KRASV12 transgenic mice, In addition, the tumors within the ApcMin KRASV12 Klf5 mice, when formed, have been smaller sized than people from your ApcMin KRASV12 mice, Without a doubt, ApcMin KRASV12 mice needed to be euthanized by 12 weeks of age, due to the presence of rectal prolapse from the large tumor burden. In contrast, nearly all ApcMin KRASV12 Klf5 mice survived as much as a 12 months without having displaying overt morbidity.
Taken into consideration that expression of selleck chemical Blebbistatin the KRASV12 transgene in the small intestine of ApcMin KRASV12 Klf5 mice remains robust, our study suggests that haploinsufficiency of Klf5 attenuates the cumulative effect of Apc inactivation and oncogenic KRAS activation. Our final results display that a mixed effect of ApcMin and KRASV12 mutations is often a major improve inside the ranges of b catenin, cyclin D1 and Ki67, within the standard appearing intestinal tissues within the ApcMin KRASV12 mice as in contrast to wild style mice, This boost is just like that noticed in the intestine in the ApcMin mice, Haploinsufficiency of Klf5 attenuated the increase while in the levels of these 3 pro teins from the normal appearing intestine of ApcMin KRASV12 mice to levels that resembled the wild sort intestine, These benefits indicate the increase in b catenin and cyclin D1 levels while in the intestine of mutant mice is largely a consequence of ApcMin mutation, instead of KRASV12 more than expression and the tumor suppressive result of Klf5 haploin sufficiency in ApcMin KRASV12 mice is due principally towards the capability of Klf5 to modulate ApcMin signaling.