Myeloproliferative Defects Following Targeting of the Drf1 Gene Encoding the Mammalian Diaphanous–Related Formin Mdia1

Priority Report

Myeloproliferative Defects following Targeting of the Drf1 Gene Encoding the Mammalian Diaphanous–Related Formin mDia1

Jun Peng,1 Susan M. Kitchen,1 Richard A. West,1,2 Robert Sigler,3 Kathryn M. Eisenmann,1 and Arthur S. Alberts1

1Laboratory of Cell Structure and Signal Integration and 2Flow Cytometry Core Facility, Van Andel Research Institute, Grand Rapids, Michigan and 3Esperion Therapeutics, Division of Pfizer, Ann Arbor, Michigan

Abstract disruption of an intramolecular mechanism maintained by conserved regulatory domains that flank the formin homology-2 Rho GTPase-effector mammalian diaphanous (mDia)–related formins assemble nonbranched actin filaments as part of domain (3, 4). cellular processes, including cell division, filopodia assembly, To date, there are only two genetic disorders associated with and intracellular trafficking.Whereas recent efforts have led the genes encoding mDia proteins (2). In the first, the DFNA1 allele to thorough characterization of formins in cytoskeletal of the DRF1/DIAPH1 (5q31) gene for human mDia1 has been remodeling and actin assembly in vitro, little is known about characterized in nonsyndromic deafness (5). The DFNA1 mutation the role of mDia proteins in vivo.To fill this knowledge gap, is likely not a loss-of-function mutation, however. The allele carries the Drf1 gene, which encodes the canonical formin mDia1, was a frameshift mutation predicted to cause a truncation near the targeted by homologous recombination.Upon birth, Drf1+/ C-terminal diaphanous autoregulatory domain (4) that may affect and Drf1 / mice were developmentally and morphologically mDia1 autoinhibition and regulation by small GTPases (6). The second DRF genetic defect is a breakpoint translocation in the indistinguishable from their wild-type littermates.However, both Drf1+/ and Drf1 / developed age-dependent myelo- last exon of the DRF2/DIAPH2 (Xq22) gene encoding human mDia3 proliferative defects.The phenotype included splenomegaly, protein; the translocation has been associated with premature fibrotic and hypercellular bone marrow, extramedullary ovarian failure in one patient (7). However, there has been no hematopoiesis in both spleen and liver, and the presence of demonstration that expression or function of mDia3 protein was immature myeloid progenitor cells with high nucleus-to- affected by this mutation. cytoplasm ratios.Analysis of cell surface markers showed an In this study of the effects of knocking out Drf1 gene expression age-dependent increase in the percentage of CD11b+-activated in mice, we show that mDia1 plays an essential role in and CD14+-activated monocytes/macrophages in both spleen myelopoiesis. As animals age, they develop myeloproliferative and bone marrow in Drf1+/ and Drf1 / animals.Analysis of defects in both the bone marrow and peripheral blood. These the erythroid compartment showed a significant increase in observations point to a crucial role of mDia1 in maintaining the proportion of splenic cells in S phase and an expansion of myeloid homeostasis, potentially by functioning as a tumor erythroid precursors (TER-119+ and CD71+)inDrf1-targeted suppressor or susceptibility gene. mice.Overall, knocking out mDia1 expression in mice leads to a phenotype similar to human myeloproliferative syndrome Materials and Methods (MPS) and myelodysplastic syndromes (MDS).These observations suggest that defective DRF1 expression or mDia1 Gene targeting, immunoblotting, and genotyping. Gene targeting, genotyping, and immunoblotting were done exactly as described in Peng function may contribute to myeloid malignancies and point et al. (8). Embryonic stem cells used in the initial targeting were from to mDia1 as an attractive therapeutic target in MDS and MPS. 129(CJ7) and chimeric mice were B6; mice used in this study were of a [Cancer Res 2007;67(16):7565–71] mixed 129/B6 background. Fluorescence-activated cell sorting analysis. Marrow, spleen, blood, and tumor cells (where applicable) from each age group and genotype were Introduction characterized by flow cytometric analyses. Marrow was flushed from femurs Formins nucleate, processively elongate, and (in some cases) using a syringe with a fine gauge needle and 3 mL of PBS. Single-cell bundle filamentous actin (F-actin) through conserved formin suspensions of spleen and tumors were obtained by mincing tissue with homology-2 domains (1). The mammalian diaphanous (mDia)– glass slides and subsequent passage and scraping of tissue in a related formins participate in many cytoskeletal remodeling events, ThermoShandon biopsy bag (Thermo Fisher Scientific). Cells were incubated for 15 min. at 20jC in the dark. Incubation was followed by including cytokinesis, vesicle trafficking, and filopodia assembly, addition of 1 FACSLyse reagent (Becton Dickinson) for 15 min at 20jCin while acting as effectors for Rho family small GTPases (2). Active the dark. After RBC lysis the remaining cells were washed in 2 mL PBS with GTP-bound Rho binds to and activates mDia proteins through 0.1% sodium azide. Cells were fixed in 1.0% methanol-free formaldehyde (Polysciences, Inc.) in PBS containing 0.1% bovine serum albumin and refrigerated at 4.0jC until acquisition. Appropriate subclass and negative controls were used to detect nonspecific binding of antibody and auto- Note: Supplementary data for this article are available at Cancer Research Online fluorescence. A minimum of 10,000 events for fresh marrow mononuclear (http://cancerres.aacrjournals.org/). cells and 5,000 events for spleen and tumor cells were acquired when Requests for reprints: Art Alberts, Research Laboratories, Van Andel Research possible. Flow cytometric analyses were conducted using either a Becton Institute, 333 Bostwick Avenue N.E., Grand Rapids, MI 49503. Phone: 616-234-5316; Dickinson FACSCalibur four-color or a FACSAria 12-color flow cytometer Fax: 616-234-5317; E-mail: [email protected]. I2007 American Association for Cancer Research. (Becton Dickinson). Data were analyzed using Becton Dickinson CellQUEST doi:10.1158/0008-5472.CAN-07-1467 ProR (v5.2.1) and FACSDiVaR (v5.0.1) software. www.aacrjournals.org 7565 Cancer Res 2007; 67: (16). August 15, 2007

Monoclonal antibodies. The following monoclonal antibodies were increased levels of monocytes and ring granulocytes. In rare cases, used: anti-CD14FITC (Sa2-8) from eBioscience; anti-CD29APC from the marrow was fibrotic, as shown in the bottom-right of Fig. 2B BioLegend; anti-CD45PerCP (30-F11), anti-CD41FITC (MWReg30), anti- (>5% of examined smears). Peripheral blood characteristics of CD71FITC (C2), anti-CD74FITC (In-1), anti-TER-119PE (Ly-76, TER-119), Drf1-targeted mice were then examined. anti-CD13PE (R3-242), anti-CD19PE (1D3), and anti-CD11bAPC (D12) from Blood smears of age-matched 450-day-old mice showed varying BD PharMingen; anti-CD8aPE (5H10), anti-CD4APC (RM4-5), anti-CD34APC (MEC14.7), and anti-CD3FITC (500A2) from Invitrogen/Caltag Laboratories. degrees of dysplasia; representative examples are shown in Fig. 2C Cell cycle analysis. Cell cycle analyses were done where applicable using compared with blood from wild-type mice. Peripheral blood smears +/+ +/ propidium iodide (Sigma) in a modified Vindelov’s preparation. Whenever from Drf1 and Drf1 were indistiguishable. However, periph- / possible, a minimum of 10,000 events were collected by flow cytometry. eral blood from age-matched Drf1 littermates was abnormal Data were analyzed using Becton Dickinson CellQUEST Pro and Verity by comparison. An example is shown in Fig. 2C (third column House ModFIT LT (v3.1) software. and inset) where abnormally shaped (dysplastic) erythrocytes, characterized as echinocytes with spiked or star-shaped appear- ance, were seen in Drf1 / samples; hemoglobin levels were within Results normal ranges (data not shown). The same animal had numerous Expression of the murine Drf1 gene encoding the mammalian immature myeloid progenitors. This was also observed in a diaphanous-related formin mDia1 (9) protein was disrupted by littermate as shown in Fig. 2C (4th column); echinocytes were homologous recombination in embryonic stem cells derived from not present in those samples. In both cases, the progenitor cells 129Sv mice as previously described (8, 10); the targeting strategy is seemed clumped with high nucleus to cytoplasm ratio. The WBC shown schematically in Fig. 1A. Drf1-targeted embryonic stem cells count in peripheral blood was abnormally elevated in this were used to generate 129Sv/B6 chimeras and the Drf1 mice were particular mouse (51.3 103/AL blood) and was significantly maintained on a B6/129Sv background. Loss of mDia1 protein elevated in a cohort of 10 Drf1 / mice compared with Drf1+/ and expression in Drf1+/ and Drf1 / mice was confirmed by Drf1+/+ age-matched littermates (Supplementary Table S1). The immunoblotting (Fig. 1A). At birth, mDia1 Drf1 / animals were high WBC levels and other hyperproliferative features in myeloid viable and morphologically normal. progenitors prompted us to examine myelopoiesis in more detail. As animals aged (>300 days of age), however, they frequently We used flow cytometric analysis to segregate bone marrow cells (30–40%) developed dermatoses and, in some cases, alopecia from 100-day-old and 450-day-old mice (n > 15, each genotype). By (Fig. 1B). H&E staining of formalin-fixed and paraffin-embedded plotting CD45 (a pan-leukocyte marker) versus side-scatter, cells skin sections routinely showed no sign of infection but high levels of segregated into discrete populations: blasts (immature cells), neutrophil invasion (inset). In addition to these outward signs erythroid precursors (EPC), lymphocytes (lymph), monocytes of ill health, animals often failed to thrive between 200 and 450 days (monos), and granulocytes (grans; as shown in the gating strategy of age and showed signs of dehydration and anemia (pale ears outlined in Fig. 3A). We found that 450-day-old Drf1+/ and Drf1 / and feet). Despite these defects, life spans of Drf1+/ and Drf1 / mice had significant expansion of cells in the granulocyte mice were not significantly different than those of their wild-type compartment (Fig. 3B). In contrast, the percentage of cells falling littermates (data not shown). into the lymph gate was diminished; these results were consistent Upon necropsy of animals found dead or euthanized at various with the lymphopenia documented in the Supplementary Table S1. ages, splenomegaly (Fig. 1C)wasobservedinbothDrf1+/ and Cells within the respective gates were then analyzed further Drf1 / mice; however, the difference in spleen mass (450 days to determine the developmental status of cells within each of age) was statistically significant only in Drf1 / animals compartment. (Fig. 1C, histogram). Histopathology revealed significant dysplasia CD14 (LPS/LBP receptor; an activation marker of monocyte and in the spleen, as shown in H&E-stained sections (Fig. 1D)from macrophage cell lineage) expression vs. CD11b (integrin aM; 100-day-old Drf1-null mice. Typically, sections showed a lack of or monocyte development marker) was assessed in spleens and bone malformed germinal centers with essentially no white pulp within marrow from mice, ages 100 and 450 days (Fig. 3C). Consistent the spleen, suggesting a defect in immune cell migration and/or with the observations shown in Fig. 3B, the percentage of cells proliferation. Similar observations were made in lymph nodes of expressing the monocyte marker CD11b remained relatively Drf1-targeted mice (data not shown). unchanged within the bone marrow across all genotypes. As Consistent with the observations in 100-day-old Drf1 / animals aged, however, a 2-fold increase in the percentage of animals, splenic sections from older Drf1+/ animals (f400 days) CD11b+ cells was observed in spleens of Drf1 / animals compared had poorly formed germinal centers but also had increased levels with wild-type animals. The percentages of CD14+ cells within either of granulocytes and neutrophils (Fig. 2A). Additional features bone marrow or spleen was enhanced in both Drf1+/ and Drf1 / included high levels of hemosidirin (a breakdown product of mice relative to wild-type counterparts. The percentages of hemoglobin), marked congestion and necrosis, and extramedullary CD14+ cells within both marrow and spleen were also increased hematopoiesis (EMH). EMH was also observed in the lung and in a time-dependent manner in Drf1 / mice. Collectively, these liver (data not shown; Fig. 2A, far right), where it was accompanied data are consistent with the notion that the activation status of by aggregates of plasma cells. the monocyte/macrophage population is enhanced upon loss of Bone marrow was examined to assess what, if any, changes mDia1 protein expression. might have occurred in the myeloid compartment. Figure 2B shows We next examined the percentages of bone marrow and splenic Giemsa-Wright–stained bone marrow obtained from f400-day-old cells—again in the lymph/mono gate—expressing the extracellular mouse femurs. Whereas Drf1+/+ mice displayed normal marrow, matrix receptor h1 integrin (CD29), as its expression is important marrow from both Drf1+/ and Drf1 / animals were markedly for both homing to and retention within lymphoid organs (Fig. 3C, hypercellular. Erythroid progenitor cells contained abnormal bottom two histograms). The percentage of cells expressing CD29 mitotic figures, abnormally high nucleus-to-cytoplasm ratios, and in marrow was not altered in either Drf1+/ or Drf1 / genotype.

Cancer Res 2007; 67: (16). August 15, 2007 7566 www.aacrjournals.org

Figure 1. Drf1 gene targeting leads to dermatoses, splenomegaly, and myelodysplasia. A, gene targeting strategy as reported previously along with example PCR-based genotyping (8). Immunoblotting for mDia1 expression in 20 Ag of whole-cell lysate using P155 rabbit anti-mDia1 (20). B, Drf1 / mice (male and female, inset; 450 d) with malocclusion (top), granulated dermatitis, and inflammation on paws. Female with dermatoses along crest and alopecia of muzzle from excessive barbering (bottom). H&E staining of formalin-fixed paraffin-embedded skin section with chronic inflammation (without any evidence of infection) and high levels of neutrophil invasion (inset). C, splenomegaly in Drf1-targeted mice (scale = cm). The histogram shows mean wet weight of spleens at necrocropsy of age-matched (450 d) mice. Columns, mean; bars, +/ standard deviation. D, H&E-stained splenic sections showing a lack of splenic nodules and T-cell infiltration sections from 450-day-old mice of the indicated genotypes. www.aacrjournals.org 7567 Cancer Res 2007; 67: (16). August 15, 2007

Figure 2. Myelodysplastic features in spleen and bone marrow. A, dysplasia in formalin-fixed H&E-stained paraffin-embedded splenic sections in 300-day-old to 450-day-old mice with indicated genotypes (described in detail in text). B, bone marrow smears from femurs of 400-day-old mice stained with Wright-Giemsa. C, Wright-Giemsa–stained peripheral blood from 400-day-old littermates.

Cancer Res 2007; 67: (16). August 15, 2007 7568 www.aacrjournals.org

Figure 3. Myeloid bone marrow composition and myeloid components of marrow. A, bone marrow composition for Drf +/+, Drf1 +/, and Drf1 / 450-day-old mice. Symbols above the columns in parts B and C (*, ., j, 1, +, **, jj, and F ) denote the paired comparisons having significant P values. P values for the comparisons are noted below. B, bone marrow shows significant differences in composition at 450 d (P < 0.01; except for ., where P < 0.05). C, comparison of the CD14+ cells from marrow and spleen at 100 and 450+ days (P < 0.01; except for . and j, where P < 0.05). There was a significant increase in CD11b+ cells in marrow; in the spleen there were significant differences in each genotype at each time point (P < 0.05; except for j, where P < 0.01). CD29 expression differences were found in marrow only in the Drf1-null relative to the other genotypes at 450+ days (P < 0.01). In spleen, there are significant differences seen at 100 d only in the null compared with the other genotypes (P < 0.05); at 450+ days there were significant differences seen in all three genotypes (P < 0.01; except for j, where P < 0.05). In both B and C, bars represent the means +/ standard deviations. www.aacrjournals.org 7569 Cancer Res 2007; 67: (16). August 15, 2007

Figure 4. Erythroid extramedullary and myeloid dysplasias. A, erythroid precursor compartment in both spleen and marrow at 450 d, where there is an increase in the number of erythroid precursors in the spleen in the Drf1-null (as in the extreme example in this figure, where 70% of the splenic tissue is erythroid precursor cells). See Fig. 3 caption for meaning of the symbols above the bars in B. B, erythroid precursors in spleen and marrow shows that the condition affects the Drf1-null animal in splenic tissue (P < 0.01). Also at 100 and 450 d, there is an increase in the splenic S phase (P < 0.01) but not in the marrow.

In contrast, the percentage of gated splenic cells expressing CD29 phenotypic aspects progressing with age. The complex Drf1 / was significantly increased in both Drf1+/ and Drf1 / mice, when phenotype includes the potential to develop neutrophilic derma- compared with wild-type littermates. This increase within the toses, splenomegaly, EMH, alopecia, and numerous cytopenias spleen was amplified as the Drf1-targeted mice aged. This and cytoses (11). Whereas the variability seen here may reflect the observation points to defects in the development and differenti- mixed genetic background of the mice, overall, the phenotype ation within the myeloid compartment of Drf1 / mice potentially resembles human chronic myeloproliferative syndromes (MPS) driving the development of splenomegaly. To further explore this and myelodysplastic syndromes (MDS). Both MPS and MDS have possibility, erythropoiesis was analyzed in Drf1-targeted mice. been characterized as preleukemic states, including variable lym- Figure 4 illustrates a comparison of TER-119 (erythroid-specific phopenia, excess or dysfunctional erythrocytes, chronic myelomo- marker) and CD71 (transferrin receptor; marker of proliferating nocytic leukemia, ineffective hematopoiesis, and, in some cases, erythroid precursors) levels in bone marrow and spleen cells from advancing myelofibrosis. There are also incidences of neutrophilic 100-day-old and 450-day-old Drf1+/ and Drf1 / animals relative dermatoses (Sweet’s syndrome) which can accompany MDS and to their wild-type counterparts. No significant differences were MPS (12). MDS is a frequent hematologic disorder that affects observed in either the relative percentages of TER-119+–gated or typically older patients and is thought to be a stem cell disorder. CD71+-gated splenic cells in 100-day-old mice (data not shown). Dysplastic features of the nucleus or cytoplasm, as observed in Yet, by 450 days of age, there was a marked elevation in the the mDia1 KO mice, and altered cellularity of the bone marrow are percentage of gated Drf1 / splenic cells expressing both CD71 also characteristic of MDS (11). and TER-119 (Fig. 4A). These data, particularly the high levels of mDia1 knockout leads to key hyperproliferation within the transferrin receptor, led us to examine whether an increase in spleen and expansion of activated monocytes/macrophages proliferation of erythroid progenitors within the spleen accounted within bone marrow and spleen. Consistent with MPS and for the observed EMH. MDS, active cell turnover and cell division was observed. MDS To test this hypothesis, the proportion of cells in S phase within and MPS give rise to clonal hematopoietic and myelopoietic bone marrow and the spleen were analyzed, as described in the defects, and whereas we have not shown clonality, we consistently Materials and Methods. As shown in Fig. 4B (right), the proportion see defects in erythropoiesis. The effect of Drf1 gene targeting of cells in S phase between different genotypes in cells from bone and the resulting mDia1 knockout suggests that the DRF1 gene marrow was unchanged at both 100 or 450 days. Consistent with for human mDia1 is affected in MPS, MDS, or other preleukemic our hypothesis, at 450 days, there was a 4-fold to 6-fold increase pathologies. in S phase in Drf1 / animals and a 2-fold to 4-fold increase in DRF1 has been mapped to 5q31.3. There are several candidate the splenic erythroid precursors from Drf1+/ mice. These data, genes in chromosome 5q in which defective expression/function in combination with the TER119 and CD71 results, suggest that could lead to myeloproliferitive defects and myelodysplasia (11). the rapidly proliferating cells within the spleen were erythroid One is the gene encoding nucleophosmin NPM1, which has been precursors. targeted in mice (13). Whereas Npm1 haploinsufficiency led to some myeloproliferative defects in mice similar to those observed here, the NPM1 (5q35) gene often falls outside of the chromo- Discussion somal abnormalities that accompany MDS (14). Another study has Knocking out mDia1 expression interferes with several aspects pointed to repressed a-catenin (CTNNA1; 5q31.2) gene expression of myelopoiesis and erythropoiesis, with the severity of certain in 5q (minus) syndrome (15); CTNNA1 has been shown to fall

Cancer Res 2007; 67: (16). August 15, 2007 7570 www.aacrjournals.org

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 2007 American Association for Cancer Research. Myeloproliferation in mDia1 Knockout Mice within a minimal region of 5q31 deleted in some forms of acute Taken as a whole, the wide range of observations suggest that myelogenous leukemia (AML)–associated with MDS (16). Whereas there are multiple mechanisms that lead to progression to CTNNA1 message is repressed, the loss of or diminished a-catenin myeloproliferative disease and progression to malignacy. The protein expression in primary patient samples has yet to be addition of the Drf1-targeted mice to this repertoire should help shown. elucidate the molecular pathogenesis of myeloproliferative and Recent evidence from carcinogen-treated mice also points to myelodysplastic disease. defects in Egr1/Krox-20 (5q31.2) expression as having a role in MDS and progression to AML (17). This finding is particularly Acknowledgments compelling because expression of Egr-1 (an immediate-early gene product) is controlled by the serum response factor (SRF; ref. 18). Received 4/20/2007; revised 6/7/2007; accepted 6/21/2007. Grant support: Van Andel Foundation, National Cancer Institute grant CA107529, Whereas Egr-1 expression is not entirely dependent upon Rho American Cancer Society grant RSG-05-033-01-CSM (A.S. Alberts, J. Peng, and S.M. GTPase activity, its expression requires SRF activation via con- Kitchen), and NRSA grant F32 GM723313 (K.M. Eisenmann). The costs of publication of this article were defrayed in part by the payment of page trolled actin dynamics (19). SRF in turn can be strongly induced charges. This article must therefore be hereby marked advertisement in accordance by activated variants of mDia1, which potently assemble F-actin in with 18 U.S.C. Section 1734 solely to indicate this fact. cells (20). Therefore, loss of mDia1 may affect Egr-1 expression, We thank David Nadziejka and Aaron DeWard for critical reading of the manuscript, Nick Duesbery for many helpful discussions, Bart Williams for critical thereby leading to similar outcomes as those observed in Egr-1– insight, and members of Miranti, MacKiegan, and Duesbery laboratories for comments targeted mice. throughout the project.

References 8. Peng J, Wallar BJ, Flanders A, Swiatek PJ, Alberts AS. syndromes and acute myeloid leukemia. Cancer Genet Disruption of the Diaphanous-related formin Drf1 gene Cytogenet 2006;1671:66–9. 1. Goode BL, Eck MJ. Mechanism and Function of encoding mDia1 reveals a role for Drf3 as an effector for 15. Liu TX, Becker MW, Jelinek J, et al. Chromosome 5q Formins in Control of Actin Assembly. Annu Rev Cdc42. Curr Biol 2003;137:534–45. deletion and epigenetic suppression of the gene Biochem 2007;76:593–627. 9. Watanabe N, Madaule P, Reid T, et al. p140mDia, a encoding a-catenin (CTNNA1) in myeloid cell transfor- 2. Wallar BJ, Alberts AS. The formins: active scaffolds that mammalian homolog of Drosophila diaphanous, is a mation. Nat Med 2007;131:78–83. remodel the cytoskeleton. Trends Cell Biol 2003;138:435–46. target protein for Rho small GTPase and is a ligand for 16. Horrigan SK, Arbieva ZH, Xie HY, et al. Delineation of 3. Watanabe N, Kato T, Fujita A, Ishizaki T, Narumiya S. profilin. EMBO J 1997;1611:3044–56. a minimal interval and identification of 9 candidates for Cooperation between mDia1 and ROCK in Rho-induced 10. Swiatek PJ, Gridley T. Perinatal lethality and defects a tumor suppressor gene in malignant myeloid dis- actin reorganization. Nat Cell Biol 1999;13:136–43. in hindbrain development in mice homozygous for a orders on 5q31. Blood 2000;957:2372–7. 4. Alberts AS. Identification of a carboxyl-terminal targeted mutation of the zinc finger gene Krox20. Genes 17. Joslin JM, Fernald AA, Tennant TR, et al. Haploinsuffi- diaphanous-related formin homology protein autoregu- Dev 1993;711:2071–84. ciency of EGR1, a candidate gene in the del(5q), leads to the latory domain. J Biol Chem 2001;2764:2824–30. 11. Corey SJ, Minden MD, Barber DL, Kantarjian H, Wang development of myeloid disorders. Blood 2007;110:719–26. 5. Lynch ED, Lee MK, Morrow JE, Welcsh PL, Leon PE, JC, Schimmer AD. Myelodysplastic syndromes: the 18. Janssen-Timmen U, Lemaire P, Mattei MG, Revelant O, King MC. Nonsyndromic deafness DFNA1 associated complexity of stem-cell diseases. Nat Rev Cancer 2007; Charnay P. Structure, chromosome mapping and regu- with mutation of a human homolog of the Drosophila 72:118–29. lation of the mouse zinc-finger gene Krox-24; evidence for gene diaphanous [see comments]. Science 1997;2785341: 12. Cohen PR, Kurzrock R. Sweet’s syndrome revisited: a a common regulatory pathway for immediate-early 1315–8. review of disease concepts. Int J Dermatol 2003;4210: serum-response genes. Gene 1989;802:325–36. 6. Higgs HN. Formin proteins: a domain-based approach. 761–78. 19. Gineitis D, Treisman R. Differential usage of signal Trends Biochem Sci 2005;306:342–53. 13. Grisendi S, Bernardi R, Rossi M, et al. Role of transduction pathways defines two types of serum response 7. Bione S, Sala C, Manzini C, et al. A human homologue of nucleophosmin in embryonic development and tumor- factor target gene. J Biol Chem 2001;27627:24531–9. the Drosophila melanogaster diaphanous gene is disrupted igenesis. Nature 2005;4377055:147–53. 20. Tominaga T, Sahai E, Chardin P, McCormick F, in a patient with premature ovarian failure: evidence for 14. Royer-Pokora B, Trost D, Muller N, Hildebrandt B, Courtneidge SA, Alberts AS. Diaphanous-related formins conserved function in oogenesis and implications for Germing U, Beier M. Delineation by molecular cytoge- bridge Rho GTPase and Src tyrosine kinase signaling. human sterility. Am J Hum Genet 1998;623:533–41. netics of 5q deletion breakpoints in myelodyplastic Molecular Cell 2000;51:13–25.

www.aacrjournals.org 7571 Cancer Res 2007; 67: (16). August 15, 2007

Downloaded from cancerres.aacrjournals.org on September 29, 2021. © 2007 American Association for Cancer Research. Myeloproliferative Defects following Targeting of the Drf1 Gene Encoding the Mammalian Diaphanous−Related Formin mDia1

Jun Peng, Susan M. Kitchen, Richard A. West, et al.

Cancer Res 2007;67:7565-7571.

Updated version Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/67/16/7565

Cited articles This article cites 20 articles, 1 of which you can access for free at: http://cancerres.aacrjournals.org/content/67/16/7565.full#ref-list-1

Citing articles This article has been cited by 23 HighWire-hosted articles. Access the articles at: http://cancerres.aacrjournals.org/content/67/16/7565.full#related-urls

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://cancerres.aacrjournals.org/content/67/16/7565. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.