Noncanonical cytoplasmic poly(A) polymerases regulate RNA levels, alternative RNA processing, and synaptic plasticity but not hippocampal-dependent behaviours.

Noncanonical poly(A) polymerases are frequently tethered to mRNA 3' untranslated regions and regulate poly(A) tail length and resulting translation. In the brain, one such poly(A) polymerase is Gld2, which is anchored to mRNA by the RNA-binding protein CPEB1 to control local translation at postsynaptic regions. Depletion of CPEB1 or Gld2 from the mouse hippocampus results in a deficit in long-term potentiation (LTP), but only depletion of CPEB1 alters animal behaviour. To test whether a related enzyme, Gld4, compensates for the lack of Gld2, we separately or simultaneously depleted both proteins from hippocampal area CA1 and again found little change in animal behaviour, but observed a deficit in LTP as well as an increase in long-term depression (LTD), two forms of protein synthesis-dependent synaptic plasticity. RNA-seq data from Gld2, Gld4, and Gld2/Gld4-depleted hippocampus show widespread changes in steady state RNA levels, alternative splicing, and alternative poly(A) site selection. Many of the RNAs subject to these alterations encode proteins that mediate synaptic function, suggesting a molecular foundation for impaired synaptic plasticity.

A role of cellular translation regulation associated with toxic Huntingtin protein.

Huntington's disease (HD) is a severe neurodegenerative disorder caused by poly Q repeat expansion in the Huntingtin (Htt) gene. While the Htt amyloid aggregates are known to affect many cellular processes, their role in translation has not been addressed. Here we report that pathogenic Htt expression causes a protein synthesis deficit in cells. We find a functional prion-like protein, the translation regulator Orb2, to be sequestered by Htt aggregates in cells. Co-expression of Orb2 can partially rescue the lethality associated with poly Q expanded Htt. These findings can be relevant for HD as human homologs of Orb2 are also sequestered by pathogenic Htt aggregates. Our work suggests that translation dysfunction is one of the contributors to the pathogenesis of HD and new therapies targeting protein synthesis pathways might help to alleviate disease symptoms.

CPSF7 regulates liver cancer growth and metastasis by facilitating WWP2-FL and targeting the WWP2/PTEN/AKT signaling pathway.

Alternative splicing within a gene can create different versions of an mRNA, called isoforms. CFIm, composed of a small subunit CFIm25 and two large subunits CFIm68 and CFIm59 (also known as CPSF7), has been proposed as an enhancer-dependent activator of mRNA 3' processing. In this study, we investigated the role of CPSF7 in hepatocellular carcinoma. Experimental evidence suggests that the expression level of CPSF7 is higher in liver cancer cells and tissues than in non-tumor hepatic cells and tissues. Furthermore, knockdown of CPSF7 effectively suppressed cell proliferation, migration and colony formation in liver cancer cells by inhibiting PTEN/AKT signaling. CPSF7 promoted WWP2-FL due to the presence of PTEN ubiquitination sites in this longer transcript. Taken together, we identified that CPSF7 regulates liver cancer growth by targeting WWP2-FL that in turn regulates AKT activation in a PTEN-dependent manner.

World Health Organization-defined eosinophilic disorders: 2019 update on diagnosis, risk stratification, and management.

DISEASE OVERVIEW: The eosinophilias encompass a broad range of non-hematologic (secondary or reactive) and hematologic (primary, clonal) disorders with potential for end-organ damage. DIAGNOSIS: Hypereosinophilia has generally been defined as a peripheral blood eosinophil count greater than 1.5 × 109 /L, and may be associated with tissue damage. After exclusion of secondary causes of eosinophilia, diagnostic evaluation of primary eosinophilias relies on a combination of various tests. They include morphologic review of the blood and marrow, standard cytogenetics, fluorescence in situ-hybridization, flow immunophenotyping, and T-cell clonality assessment to detect histopathologic or clonal evidence for an acute or chronic hematolymphoid neoplasm. RISK STRATIFICATION: Disease prognosis relies on identifying the subtype of eosinophilia. After evaluation of secondary causes of eosinophilia, the 2016 World Health Organization endorses a semi-molecular classification scheme of disease subtypes. This includes the major category "myeloid/lymphoid neoplasms with eosinophilia and rearrangement of PDGFRA, PDGFRB, or FGFR1 or with PCM1-JAK2", and the MPN subtype, "chronic eosinophilic leukemia, not otherwise specified" (CEL, NOS). Lymphocyte-variant hypereosinophilia is an aberrant T-cell clone-driven reactive eosinophila, and idiopathic hypereosinophilic syndrome (HES) is a diagnosis of exclusion. RISK-ADAPTED THERAPY: The goal of therapy is to mitigate eosinophil-mediated organ damage. For patients with milder forms of eosinophilia (eg, <1.5 × 109 /L) without symptoms or signs of organ involvement, a watch and wait approach with close-follow-up may be undertaken. Identification of rearranged PDGFRA or PDGFRB is critical because of the exquisite responsiveness of these diseases to imatinib. Corticosteroids are first-line therapy for patients with lymphocyte-variant hypereosinophilia and HES. Hydroxyurea and interferon-alfa have demonstrated efficacy as initial treatment and in steroid-refractory cases of HES. In addition to hydroxyurea, second line cytotoxic chemotherapy agents, and hematopoietic stem cell transplantation have been used for aggressive forms of HES and CEL, with outcomes reported for limited numbers of patients. The use of antibodies against interleukin-5 (IL-5) (mepolizumab), the IL-5 receptor (benralizumab), as well as other targets on eosinophils remains an active area of investigation.

The Influenza A Virus Endoribonuclease PA-X Usurps Host mRNA Processing Machinery to Limit Host Gene Expression.

Many viruses shut off host gene expression to inhibit antiviral responses. Viral proteins and host proteins required for viral replication are typically spared in this process, but the mechanisms of target selectivity during host shutoff remain poorly understood. Using transcriptome-wide and targeted reporter experiments, we demonstrate that the influenza A virus endoribonuclease PA-X usurps RNA splicing to selectively target host RNAs for destruction. Proximity-labeling proteomics reveals that PA-X interacts with cellular RNA processing proteins, some of which are partially required for host shutoff. Thus, PA-X taps into host nuclear pre-mRNA processing mechanisms to destroy nascent mRNAs shortly after their synthesis. This mechanism sets PA-X apart from other viral host shutoff proteins that target actively translating mRNAs in the cytoplasm. Our study reveals a unique mechanism of host shutoff that helps us understand how influenza viruses suppress host gene expression.

Pemphigus-like hypereosinophilic syndrome with FIP1L1-PDGFRA fusion gene: A challenging and uncommon clinical presentation.

Hypereosinophilic syndrome (HES) is often associated with cutaneous manifestations, mostly pruritic lesions, urticaria and angioedema. Mucosal lesions are rarely seen in HES but, when present, are usually the first manifestation of the disease. The clinical presentation may be heterogeneous, including erosions, aphthae or ulcers, and can be easily confused with other mucocutaneous disorders. Here, we present the case of a 64-year-old man with severe chronic erosive oral mucositis simulating pemphigus in which the finding of persistent eosinophilia and elevation of B12 vitamin serum levels raised the suspicion of HES. The FIP1L1-PDGFRA fusion gene (4q12) was detected by fluorescence in situ hybridization and the patient was treated with imatinib mesylate with complete response of the disease.

Targeting the polyadenylation factor EhCFIm25 with RNA aptamers controls survival in Entamoeba histolytica.

Messenger RNA 3'-end polyadenylation is an important regulator of gene expression in eukaryotic cells. In our search for new ways of treating parasitic infectious diseases, we looked at whether or not alterations in polyadenylation might control the survival of Entamoeba histolytica (the agent of amoebiasis in humans). We used molecular biology and computational tools to characterize the mRNA cleavage factor EhCFIm25, which is essential for polyadenylation in E. histolytica. By using a strategy based on the systematic evolution of ligands by exponential enrichment, we identified single-stranded RNA aptamers that target EhCFIm25. The results of RNA-protein binding assays showed that EhCFIm25 binds to the GUUG motif in vitro, which differs from the UGUA motif bound by the homologous human protein. Accordingly, docking experiments and molecular dynamic simulations confirmed that interaction with GUUG stabilizes EhCFIm25. Incubating E. histolytica trophozoites with selected aptamers inhibited parasite proliferation and rapidly led to cell death. Overall, our data indicate that targeting EhCFIm25 is an effective way of limiting the growth of E. histolytica in vitro. The present study is the first to have highlighted the potential value of RNA aptamers for controlling this human pathogen.

An effective and efficient method of transfecting primary human chondrocytes in suspension.

Human chondrocytes accumulate an ECM-rich matrix by secreting matrix macromolecules during monolayer culture, which makes them difficult to transfect efficiently. Here we report a non-viral based protocol to transfect the primary human chondrocytes with high efficiency in suspension. Chondrocyte cultures were digested using Pronase and Collagenase and transfected in suspension. Transfection efficiencies of more than 80% were achieved routinely using the protocol described. The viability of siRNA transfected or un-transfected chondrocytes was not affected and resulted in 80-90% knockdown of the target mRNA levels. This protocol may be useful in gene knockdown, and ectopic overexpression studies in chondrocytes.

Management of Hypereosinophilic Syndromes.

The symptomatic hypereosinophilic patient must be approached in a stepwise manner, with thorough assessment to determine whether the hypereosinophilia itself is contributing to damage and disease manifestations (thereby defining a hypereosinophilic syndrome), and to identify an eventual cause of hypereosinophilia, followed by initiation of treatment directed against the underlying condition or deleterious hypereosinophilic state. Situations encountered in the clinic are extremely heterogeneous because of the numerous potential causes of hypereosinophilia and the variable spectrum of eosinophil-mediated organ damage. A practical approach to many of these situations is presented in this review.

BI-2 destabilizes HIV-1 cores during infection and Prevents Binding of CPSF6 to the HIV-1 Capsid.

BACKGROUND: The recently discovered small-molecule BI-2 potently blocks HIV-1 infection. BI-2 binds to the N-terminal domain of HIV-1 capsid. BI-2 utilizes the same capsid pocket used by the small molecule PF74. Although both drugs bind to the same pocket, it has been proposed that BI-2 uses a different mechanism to block HIV-1 infection when compared to PF74. FINDINGS: This work demonstrates that BI-2 destabilizes the HIV-1 core during infection, and prevents the binding of the cellular factor CPSF6 to the HIV-1 core. CONCLUSIONS: Overall this short-form paper suggests that BI-2 is using a similar mechanism to the one used by PF74 to block HIV-1 infection.

Meiotic long non-coding meiRNA accumulates as a dot at its genetic locus facilitated by Mmi1 and plays as a decoy to lure Mmi1.

Long non-coding RNAs (lncRNAs) play key roles in the formation of nuclear bodies. In the fission yeast Schizosaccharomyces pombe, a lncRNA species termed meiRNA forms a nuclear dot structure at its own genetic locus, the sme2 locus, with its protein-binding partner Mei2. This dot structure, called Mei2 dot, promotes the progression of meiosis by suppressing Mmi1, a crucial factor involved in the selective elimination of meiosis-specific transcripts. The meiRNA itself is a target of Mmi1-mediated elimination and is supposed to function as a decoy to lure Mmi1. However, detailed mechanisms underlying the formation of Mei2 dot and inactivation of Mmi1 remain ambiguous. Here, we show that the localization of meiRNA, at its genetic locus sme2, depends on its association with Mmi1. We also demonstrate that one of the multiple Mmi1 foci in mitotic cells localizes to the sme2 locus. Furthermore, the overexpression of meiRNA promotes the accumulation of Mmi1 to the sme2 locus even in the absence of Mei2 and reduces the activity of Mmi1. These findings indicate that the retention of meiRNA at its genetic locus is facilitated by Mmi1, which then attracts scattered Mmi1 to inhibit its function.

TNPO3 protects HIV-1 replication from CPSF6-mediated capsid stabilization in the host cell cytoplasm.

BACKGROUND: Despite intensive investigation the mechanism by which HIV-1 reaches the host cell nucleus is unknown. TNPO3, a karyopherin mediating nuclear entry of SR-proteins, was shown to be required for HIV-1 infectivity. Some investigators have reported that TNPO3 promotes HIV-1 nuclear import, as would be expected for a karyopherin. Yet, an equal number of investigators have failed to obtain evidence that supports this model. Here, a series of experiments were performed to better elucidate the mechanism by which TNPO3 promotes HIV-1 infectivity. RESULTS: To examine the role of TNPO3 in HIV-1 replication, the 2-LTR circles that are commonly used as a marker for HIV-1 nuclear entry were cloned after infection of TNPO3 knockdown cells. Potential explanation for the discrepancy in the literature concerning the effect of TNPO3 was provided by sequencing hundreds of these clones: a significant fraction resulted from autointegration into sites near the LTRs and therefore were not bona fide 2-LTR circles. In response to this finding, new techniques were developed to monitor HIV-1 cDNA, including qPCR reactions that distinguish 2-LTR circles from autointegrants, as well as massive parallel sequencing of HIV-1 cDNA. With these assays, TNPO3 knockdown was found to reduce the levels of 2-LTR circles. This finding was puzzling, though, since previous work has shown that the HIV-1 determinant for TNPO3-dependence is capsid (CA), an HIV-1 protein that forms a mega-dalton protein lattice in the cytoplasm. TNPO3 imports cellular splicing factors via their SR-domain. Attention was therefore directed towards CPSF6, an SR-protein that binds HIV-1 CA and inhibits HIV-1 nuclear import when the C-terminal SR-domain is deleted. The effect of 27 HIV-1 capsid mutants on sensitivity to TNPO3 knockdown was then found to correlate strongly with sensitivity to inhibition by a C-terminal deletion mutant of CPSF6 (R2 = 0.883, p < 0.0001). TNPO3 knockdown was then shown to cause CPSF6 to accumulate in the cytoplasm. Mislocalization of CPSF6 to the cytoplasm, whether by TNPO3 knockdown, deletion of the CPSF6 nuclear localization signal, or by fusion of CPSF6 to a nuclear export signal, resulted in inhibition of HIV-1 replication. Additionally, targeting CPSF6 to the nucleus by fusion to a heterologous nuclear localization signal rescued HIV-1 from the inhibitory effects of TNPO3 knockdown. Finally, mislocalization of CPSF6 to the cytoplasm was associated with abnormal stabilization of the HIV-1 CA core. CONCLUSION: TNPO3 promotes HIV-1 infectivity indirectly, by shifting the CA-binding protein CPSF6 to the nucleus, thus preventing the excessive HIV-1 CA stability that would otherwise result from cytoplasmic accumulation of CPSF6.

Translational control of mitochondrial energy production mediates neuron morphogenesis.

Mitochondrial energy production is a tightly regulated process involving the coordinated transcription of several genes, catalysis of a plethora of posttranslational modifications, and the formation of very large molecular supercomplexes. The regulation of mitochondrial activity is particularly important for the brain, which is a high-energy-consuming organ that depends on oxidative phosphorylation to generate ATP. Here we show that brain mitochondrial ATP production is controlled by the cytoplasmic polyadenylation-induced translation of an mRNA encoding NDUFV2, a key mitochondrial protein. Knockout mice lacking the Cytoplasmic Polyadenylation Element Binding protein 1 (CPEB1) have brain-specific dysfunctional mitochondria and reduced ATP levels, which is due to defective polyadenylation-induced translation of electron transport chain complex I protein NDUFV2 mRNA. This reduced ATP results in defective dendrite morphogenesis of hippocampal neurons both in vitro and in vivo. These and other results demonstrate that CPEB1 control of mitochondrial activity is essential for normal brain development.

Cyclin-dependent kinase 7/9 inhibitor SNS-032 abrogates FIP1-like-1 platelet-derived growth factor receptor α and bcr-abl oncogene addiction in malignant hematologic cells.

PURPOSE: The "gate-keeper" mutations T674I platelet-derived growth factor receptor α (PDGFRα) in hypereosinophilic syndrome (HES) and T315I Bcr-Abl in chronic myeloid leukemia (CML) are resistant to imatinib and the second-generation small-molecule tyrosine kinase inhibitors (TKI). However, to combat acquired resistance to imatinib, an alternative approach is to decrease the expression of the addicted gene to efficiently kill resistant malignant hematologic cells. The purpose of this study was to evaluate the strategy of shutting down the transcription and expression of FIP1-like-1 (FIP1L1)-PDGFRα and Bcr-Abl with SNS-032, an inhibitor of cyclin-dependent kinase 7 (CDK7) and CDK9 in phase I clinical trials. EXPERIMENTAL DESIGN: The effects of SNS-032 on PDGFRα and Bcr-Abl signaling pathways, apoptosis, and cell cycling were analyzed in TKI-resistant cells of HES and CML. The in vivo antitumor activity of SNS-032 was assessed with xenografted BaF3-T674I FIP1L1-PDGFRα and KBM5-T315I Bcr-Abl cells in nude mouse models. RESULTS: SNS-032 inhibited the phosphorylation on Ser5 and Ser2 of RNA polymerase II. SNS-032 decreased both the mRNA and protein levels of FIP1L1-PDGFRα and Bcr-Abl and inhibited the proliferation of malignant cells expressing FIP1L1-PDGFRα or Bcr-Abl. It also decreased the phosphorylation of downstream molecules. It induced apoptosis by triggering both the mitochondrial pathway and the death receptor pathway. CONCLUSIONS: This CDK7/9 inhibitor potently inhibits FIP1L1-PDGFRα-positive HES cells and Bcr-Abl-positive CML cells regardless of their sensitivity to imatinib. SNS-032 may have potential in treating hematologic malignancy by abrogating oncogene addiction.

Pentatricopeptide repeat proteins stimulate mRNA adenylation/uridylation to activate mitochondrial translation in trypanosomes.

The majority of trypanosomal mitochondrial pre-mRNAs undergo massive uridine insertion/deletion editing, which creates open reading frames. Although the pre-editing addition of short 3' A tails is known to stabilize transcripts during and after the editing, the processing event committing the fully edited mRNAs to translation remained unknown. Here, we show that a heterodimer of pentatricopeptide repeat-containing (PPR) proteins, termed kinetoplast polyadenylation/uridylation factors (KPAFs) 1 and 2, induces the postediting addition of A/U heteropolymers by KPAP1 poly(A) polymerase and RET1 terminal uridyltransferase. Edited transcripts bearing 200- to 300-nucleotide-long A/U tails, but not short A tails, were enriched in translating ribosomal complexes and affinity-purified ribosomal particles. KPAF1 repression led to a selective loss of A/U-tailed mRNAs and concomitant inhibition of protein synthesis. These results establish A/U extensions as the defining cis-elements of translation-competent mRNAs. Furthermore, we demonstrate that A/U-tailed mRNA preferentially interacts with the small ribosomal subunit, whereas edited substrates and complexes bind to the large subunit.

Triptolide abrogates oncogene FIP1L1-PDGFRalpha addiction and induces apoptosis in hypereosinophilic syndrome.

The pathogenesis of hypereosinophilic syndrome (HES) in some patients is highly dependent on FIP1-Like-1 (FIP1L1)-platelet-derived growth factor receptor alpha (PDGFRalpha), which can generate sustained activation signaling to maintain a cell malignant phenotype. HES usually shows good response to the tyrosine kinase inhibitor imatinib, but mutations in FIP1L1-PDGFRalpha (e.g. T674I) can confer acquired resistance to imatinib. An alternative therapeutic strategy other than with tyrosine kinase inhibitors is needed to overcome acquired drug resistance. We hypothesized that switching off the crucial chimeric oncoprotein FIP1L1-PDGFRalpha on which HES cells depend, should have deleterious effects on the cancer cells. We used low concentrations of triptolide, a transcription inhibitor, to shut down the expression of FIP1L1-PDGFRalpha. EOL-1 cells and BaF3 cells expressing wild-type or T674I FIP1L1-PDGFRalpha were treated with triptolide, and signaling pathways, cell cycling, and apoptosis were analyzed by RT-PCR, immunoblotting, and flow cytometry, respectively. The results revealed that at nanomolar concentrations triptolide decreased the levels of mRNA and protein of FIP1L1-PDGFRalpha and the growth of the neoplastic cells, regardless of the mutational status of PDGFRalpha. Triptolide also downregulated the signaling molecules Stat3, Akt, and Erk1/2, which are downstream from PDGFRalpha, and induced G1 cell-cycle arrest. Triptolide time- and dose-dependently induced apoptosis by decreasing the anti-apoptotic proteins Mcl-1 and Bcl-X(L),triggering the intrinsic apoptotic pathway. In conclusion, triptolide has potent activity against malignant cells in HES bearing FIP1L1-PDGFRalpha, regardless of its mutational status that confer acquired resistance to imatinib. Our results suggest that triptolide may be a promising agent in the treatment of HES.

Inactivation of cleavage factor I components Rna14p and Rna15p induces sequestration of small nucleolar ribonucleoproteins at discrete sites in the nucleus.

Small nucleolar RNAs (snoRNAs) associate with specific proteins forming small nucleolar ribonucleoprotein (snoRNP) particles, which are essential for ribosome biogenesis. The snoRNAs are transcribed, processed, and assembled in snoRNPs in the nucleoplasm. Mature particles are then transported to the nucleolus. In yeast, 3'-end maturation of snoRNAs involves the activity of Rnt1p endonuclease and cleavage factor IA (CFIA). We report that after inhibition of CFIA components Rna14p and Rna15p, the snoRNP proteins Nop1p, Nop58p, and Gar1p delocalize from the nucleolus and accumulate in discrete nucleoplasmic foci. The U14 snoRNA, but not U3 snoRNA, similarly redistributes from the nucleolus to the nucleoplasmic foci. Simultaneous depletion of either Rna14p or Rna15p and the nuclear exosome component Rrp6p induces accumulation of poly(A)(+) RNA at the snoRNP-containing foci. We propose that the foci detected after CFIA inactivation correspond to quality control centers in the nucleoplasm.

Human Pcf11 enhances degradation of RNA polymerase II-associated nascent RNA and transcriptional termination.

The poly(A) (pA) signal possesses a dual function in 3' end processing of pre-mRNA and in transcriptional termination of RNA polymerase II (Pol II) for most eukaryotic protein-coding genes. A key protein factor in yeast and Drosophila Pol II transcriptional termination is the 3'-end processing factor, Pcf11. In vitro studies suggest that Pcf11 is capable of promoting the dissociation of Pol II elongation complexes from DNA. Moreover, several mutant alleles of yeast Pcf11 effect termination in vivo. However, functions of human Pcf11 (hPcf11) in Pol II termination have not been explored. Here we show that depletion of hPcf11 from HeLa cells reduces termination efficiency. Furthermore, we provide evidence that hPcf11 is required for the efficient degradation of the 3' product of pA site cleavage. Finally, we show that these functions of hPcf11 require an intact pA signal.

GW body disassembly triggered by siRNAs independently of their silencing activity.

GW bodies (or P-bodies) are cytoplasmic granules containing proteins involved in both mRNA degradation and storage, including the RNA interference machinery. Their mechanism of assembly and function are still poorly known although their number depends upon the flux of mRNA to be stored or degraded. We show here that silencing of the translational regulator CPEB1 leads to their disappearance, as reported for other GW body components. Surprisingly, the same results were obtained with several siRNAs targeting genes encoding proteins unrelated to mRNA metabolism. The disappearance of GW bodies did not correlate with the silencing activity of the siRNA and did not inhibit further silencing by siRNA. Importantly, in most cases, GW bodies were rapidly reinduced by arsenite, indicating that their assembly was not prevented by the inhibition of the targeted or off-target genes. We therefore propose that some siRNA sequences affect mRNA metabolism so as to diminish the amount of mRNA directed to the GW bodies. As an exception, GW bodies were not reinduced following Rck/p54 depletion by interference, indicating that this component is truly required for the GW body assembly. Noteworthy, Rck/p54 was dispensable for the assembly of stress granules, in spite of their close relationship with the GW bodies.

Molecular mechanisms underlying FIP1L1-PDGFRA-mediated myeloproliferation.

An interstitial deletion on chromosome 4q12 resulting in the formation of the FIP1L1-PDGFRA fusion protein is involved in the pathogenesis of imatinib-sensitive chronic eosinophilic leukemia. The molecular mechanisms underlying the development of disease are largely undefined. Human CD34(+) hematopoietic progenitor cells were used to investigate the role of FIP1L1-PDGFRA in modulating lineage development. FIP1L1-PDGFRA induced both proliferation and differentiation of eosinophils, neutrophils, and erythrocytes in the absence of cytokines, which could be inhibited by imatinib. Whereas expression of FIP1L1-PDGFRA in hematopoietic stem cells and common myeloid progenitors induced the formation of multiple myeloid lineages, expression in granulocyte-macrophage progenitors induced only the development of eosinophils, neutrophils, and myeloblasts. Deletion of amino acids 30 to 233 in the FIP1L1 gene [FIP1L1(1-29)-PDGFRA] gave rise to an intermediate phenotype, exhibiting a dramatic reduction in the number of erythrocytes. FIP1L1-PDGFRA and FIP1L1(1-29)-PDGFRA both induced the activation of p38 and extracellular signal-regulated kinase 1/2 (ERK1/2) in myeloid progenitors, whereas signal transducers and activators of transcription 5 (STAT5) and protein kinase B/c-akt were only activated by FIP1L1-PDGFRA. Dominant-negative STAT5 partially inhibited FIP1L1-PDGFRA-induced colony formation, whereas combined inhibition of phosphatidylinositol-3-kinase and ERK1/2 significantly reversed FIP1L1-PDGFRA-induced colony formation. Taken together, these results suggest that expression of FIP1L1-PDFGRA in human hematopoietic progenitors induce a myeloproliferative phenotype via activation of multiple signaling molecules including phosphatidylinositol-3-kinase, ERK1/2, and STAT5.