IntroductionAcute myeloid leukemia (AML) is an aggressive form of a malignant disorder of the hematopoietic system that shows increasing incidence with age and is characterized by highly proliferative blast cells. [1][2][3] Aberrant activity of tyrosine kinases has been shown to be present in various malignant diseases and AML is no exception. The FMS-like tyrosine kinase 3 (FLT3) gene in chromosome band 13q12 encodes a receptor tyrosine kinase that belongs to the same family as FMS, KIT, and the 2 genes encoding PDGFR␣ and ␤. [4][5][6][7] It is normally expressed by hematopoietic stem/progenitor cells (HSPCs) and as hematopoietic cells differentiate FLT3 expression is lost. [8][9][10][11] A large body of work has shown that FLT3 plays roles in survival, proliferation, and differentiation. Aside from its role in regulating normal hematopoiesis, FLT3 is also highly expressed in several hematologic malignancies. 7 Mutations in the receptor, in the form of internal tandem duplication (ITD) of the juxtamembrane domain and point mutations of the kinase domain, both result in constitutive activation. 7 These mutations occur in one third of AML patients making it one of the most commonly mutated genes in AML. 12,13 Patients with FLT3/ITD mutations have been demonstrated to have very poor prognosis. 5,14,15 However, the molecular basis by which FLT3/ITD mutations lead to aggressive disease and poor prognosis in AML is not yet clearly understood.The process of activation, internalization, and degradation of FLT3 occurs in a similar fashion to other members of the class III receptor-type tyrosine kinases (RTKs) family. 16 Binding of FLT3 ligand (FL) causes homodimerization, tyrosine kinase activation, receptor autophosphorylation, and initiation of downstream signaling cascades. 7 The FLT3 receptor kinase shares structural homology with other type III receptor kinases, such as KIT and FMS, with all 3 playing an important role in survival, proliferation, and differentiation of hematopoietic cells. 17 We and other investigators have shown that STAT5 is one of the principal pathways involved in mediating gene expression in response to constitutive receptor activation through mutation. FLT3 signaling results in activation of pathways through phosphorylation of STAT5, MAPK, AKT, VAV, CBL, and BAD. [18][19][20] Phosphorylation and activation of STAT5 by FLT3/ITD mutants are particularly strong compared with its phosphorylation in the wild-type FLT3 allele. 21 STAT5 tyrosine phosphorylation mediates STAT5 protein dimerization through a mechanism involving SH2 domains and N-terminal regions and results in translocation to the nucleus where it activates transcription of a number of genes. 22 However, recent data suggest that STAT5 may have other interacting partners. 23,24 For example, signal-transducing adapter proteins (STAPs) have been shown to constitutively interact with inactive STAT5 in the cytoplasm but dissociate when STAT5 is phosphorylated. 24 STAT5 is also phosphorylated on serine residues, possibly by the ERK1/2 prote...

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In this report, we investigate the role of the RNA-binding protein HuR during skeletal myogenesis. At the onset of myogenesis in differentiating C2C12 myocytes and in vivo in regenerating mouse muscle, HuR cytoplasmic abundance increased dramatically, returning to a predominantly nuclear presence upon completion of myogenesis. mRNAs encoding key regulators of myogenesis-specific transcription (myogenin and MyoD) and cell cycle withdrawal (p21), bearing AU-rich regions, were found to be targets of HuR in a differentiationdependent manner. Accordingly, mRNA half-lives were highest during differentiation, declining when differentiation was completed. Importantly, HuR-overexpressing C2C12 cells displayed increased target mRNA expression and half-life and underwent precocious differentiation. Our findings underscore a critical function for HuR during skeletal myogenesis linked to HuR's coordinate regulation of muscle differentiation genes.

The ends of linear chromosomes are capped by protein–DNA complexes termed telomeres. Telomere repeat binding factors 1 and 2 (TRF1 and TRF2) bind specifically to duplex telomeric DNA and are critical components of functional telomeres. Consequences of telomere dysfunction include genomic instability, cellular apoptosis or senescence and organismal aging. Mild oxidative stress induces increased erosion and loss of telomeric DNA in human fibroblasts. We performed binding assays to determine whether oxidative DNA damage in telomeric DNA alters the binding activity of TRF1 and TRF2 proteins. Here, we report that a single 8-oxo-guanine lesion in a defined telomeric substrate reduced the percentage of bound TRF1 and TRF2 proteins by at least 50%, compared with undamaged telomeric DNA. More dramatic effects on TRF1 and TRF2 binding were observed with multiple 8-oxo-guanine lesions in the tandem telomeric repeats. Binding was likewise disrupted when certain intermediates of base excision repair were present within the telomeric tract, namely abasic sites or single nucleotide gaps. These studies indicate that oxidative DNA damage may exert deleterious effects on telomeres by disrupting the association of telomere-maintenance proteins TRF1 and TRF2.

While transport of RNA-binding protein HuR from nucleus to cytoplasm is emerging as a key regulatory step for HuR function, the mechanisms underlying this process remain poorly understood. Here, we report that the AMP-activated kinase (AMPK), an enzyme involved in responding to metabolic stresses, potently regulates the levels of cytoplasmic HuR. Inhibition of AMPK, accomplished either through cell treatment or by adenovirus infection to express dominant-negative AMPK, was found to increase the level of HuR in the cytoplasm and to enhance the binding of HuR to p21, cyclin B1, and cyclin A mRNA transcripts and elevate their expression and half-lives. Conversely, AMPK activation, achieved by means including infection to express constitutively active AMPK, resulted in reduced cytoplasmic HuR; decreased levels and half-lives of mRNAs encoding p21, cyclin A, and cyclin B1; and diminished HuR association with the corresponding transcripts. We therefore propose a novel function for AMPK as a regulator of cytoplasmic HuR levels, which in turn influences the mRNAstabilizing function of HuR and the expression of HuR target transcripts.