Dicistronic MLV-retroviral vectors transduce neural precursors in vivoand co-express two genes in their differentiated neuronal progeny
© Derrington et al; licensee BioMed Central Ltd. 2005
Received: 07 July 2005
Accepted: 29 September 2005
Published: 29 September 2005
Dicistronic MLV-based retroviral vectors, in which two IRESes independently initiate the translation of two proteins from a single RNA, have been shown to direct co-expression of proteins in several cell culture systems. Here we report that these dicistronic retroviral vectors can drive co-expression of two gene products in brain cells in vivo. Injection of retroviral vector producer cells leads to the transduction of proliferating precursors in the external granular layer of the cerebellum and throughout the ventricular regions. Differentiated neurons co-expressing both transgenes were observed in the cerebellum and in lower numbers in distant brain regions such as the cortex. Thus, we describe an eukaryotic dicistronic vector system that is capable of transducing mouse neural precursors in vivo and maintaining the expression of genes after cell differentiation.
The brain constitutes one of the most important organs for gene therapy. Considerable interest resides in the development of vector-based therapies for many of the brain diseases, either to allow the expression of exogenous genes to compensate for a metabolic deficit, to express a growth factor and thus inhibit neural degeneration or to target suicide genes to cancer cells. An alternative approach has been the development of cellular vectors [1–3]. Uncommitted neural precursor cells can be isolated, transduced and grafted into host brains. They adapt to novel environments by stable integration and the expression of location-appropriate phenotypes in host. This opens new avenues for the use of neural stem cells as cellular vectors for gene therapy in the central nervous system (CNS) [2–6]. Both endogenous and transplanted stem cells spontaneously migrate to the site of lesions where they integrate to repopulate the damaged tissue [7–9].
Retroviral vectors based on the γ-retrovirus murine leukemia virus (MLV) are of particular interest for the transduction of neural precursor cells either ex vivo to generate cellular vectors, or in vivo to directly target endogenous neural precursors. They specifically target proliferating cells , integrate into the host genome and are conserved in cellular progeny . This has made MLV-vectors a tool of choice to trace lineages and assess the function of specific genes in rodent CNS in vivo [12–14]. In previous studies we established that dicistronic MLV-based retroviral vectors efficiently transduced cells derived from a transformed human neural stem cell line , or cells from a primary culture of neural precursors . Here we report that dicistronic MLV-based vectors can deliver and maintain expression of marker genes during neural differentiation in the CNS of new born mice.
Vector producer cells were injected in the region of the developing cerebellum, where the generation of neurons from proliferating precursors continues after birth [17–21]. At early periods post-injection, transduced cells were observed in the external granular layer (EGL) of precursors and migrating towards the internal granular layer (IGL). At later time differentiated neurons were observed scattered about the IGL or in patches. Analysis of other brain regions demonstrated a large number of transduced cells in the ependymal walls throughout the ventricular system and in the subventricular zone. Thus, our results show that dicistronic MLV-based vectors co-expressing two marker transgenes, human placental alkaline phosphatase (PLAP) and neomycin phosphotransferase (Neo) [22, 23], transduce proliferating neural precursors in vivo and can penetrate throughout the ventricular system when producer cells are grafted to host animals. Moreover, transduced neural precursors maintain expression of both transgenes after differentiation into neurons demonstrating that the activity of both internal ribosome entry segments (IRES) used in their design is not altered in vivo by neural differentiation.
Location of MLV-vector producer cells
Transgene expression in differentiated cerebellar cells in vivo
Taken together these results show that MLV-IRES vectors are able to transduce precursor cells in the CNS in vivo and that the IRESes of different viruses such as MLV, EMCV and REV-A remain functional in differentiated neurons in the animal.
Transduction of cells in different brain regions
Several neural precursor cell populations may be susceptible to transduction by the dicistronic MLV vectors in the ventricular zone. Ependymal cells, which have been reported to be neural precursors , are still proliferating quite quickly in early postnatal brain . Radial glia, which may be precursors of both neurons and glia [31–35], contact the ventricular surface and proliferate in the ventricular region until postnatal day 7 . Lastly, the slowly proliferating GFAP-labeled subependymal neural stem cells, which survive and continue to proliferate and generate neurons in the adult brain, have been proposed to require contact with the ventricular surface to become neurogenic . Having identified this sub-population of potential precursor cells we sought cells with mature phenotypes that may represent their progeny. In the most superficial layers of the cortex, which will be formed from the latest neurogenerative mitoses of cortical precursor cells, we were surprised to find rare transduced neurons, co-expressing PLAP and HU (Fig. 6H–I). Other non-neuronal cells labeled with the transgenes were also observed in forebrain (Fig. 6H,I,J and 6K).
These results showed that rather large numbers of transduced non-neuronal cells were found in and adjacent to the ependymal walls throughout the ventricular system including in the lateral ventricles (Fig. 6) and that the viral IRESes were active in these cells in vivo.
MLV-based double-IRES vectors pREV-HW3 and pEMCV-CBT4 were found to direct co-expression of two gene products in a variety of cell types [15, 16, 22, 23]. Overall transgene expression driven from the MLV-based double-IRES vectors is the consequence of two distinct processes, transcription and translation initiation, both of which are tightly regulated by the host cell. Indeed, important limitations of MLV-vectors are cell-type-specific promoter silencing [13, 37, 38], and modulation of IRES activity. IRES activity can be regulated by diverse physiological processes such as cell cycle [39–42], cellular stress [43–46], cell transformation , cell death [48–51] and cell differentiation [52–56]. Previous studies suggested that the activity of the MLV IRES present in both vectors, could be modulated by oligodendrocyte differentiation . The possibility of in vivo IRES regulation due to cell differentiation prompted us to extend our previous ex vivo studies [15, 16], and evaluate the feasibility of using double-IRES MLV vectors in the CNS.
Results show that upon injection of producer cells in the cerebellum of newborn mice, the generated MLV-vectors transduce host cells throughout the postnatal brain ventricular system. Transduced neural precursors and their progeny could be revealed by histochemistry for PLAP or immunohistochemistry and large patches of transduced neurons could be identified expressing both transgenes 15 dpi. In double labeling studies, most cells that were PLAP positive also stained for Neo. Considering that MLV-vector producer cells appeared to survive less than 10 days in the developing brain these observations demonstrated transduction of proliferating precursor cells and the maintenance of IRES activity in neurons with each of the combinations of IRESes tested, namely MLV and REV-A (pHW3) and MLV and EMCV (pCBT4). Therefore, and consistent with previous observations [15, 16], down-regulation of transgene expression was not observed in neurons generated from precursors transduced in vivo.
Transduced cells could be identified in the ependymal walls throughout the ventricular system. The 3rd and lateral ventricles lie upstream of the injection site with respect to the flow of cerebrospinal fluid. Thus, the MLV vector is capable of diffusing via the cerebrospinal fluid and targets proliferating cells in this region of the brain. Diverse cell populations identified as sources of neuronal and glial precursors are potential targets for MLV-recombinant vector in early postnatal brain [27–29, 34, 35]. For example, the proliferation of ependymal cells, which form the interface between the CSF and the brain parenchyma, progressively slows down during postnatal development to a very low basal level at postnatal day 12 which then remains stable in the absence of injury [30, 57]. Mitotic radial glia are also in contact with the ventricular surface in the lateral ventricles during early postnatal development . Subventricular astrocytes also contact the ventricular surface in adult brain and proliferate slowly .
In the ventricular regions the majority of transduced cells express GFAP, which is is weakly expressed by ependymal cells and tanycytes and more strongly expressed by astrocytes and the GFAP labeled "type B" cells that constitute multipotent neural precursors [28, 58]. Radial glia are also GFAP positive . 15 dpi, transduced cells were observed in the brain parenchyma close to the ependymal wall. In the more superficial layers of the cortex, where the most immature post mitotic neurons reside, a few transduced neurons, identified by their expression of the HU antigen, as well as non-neuronal cells could be identified.
The current approaches for gene therapy of monogenetic diseases in mature organisms are confronted by several problems including: (1) adult tissues may be poorly infected by conventional vector systems dependent upon cell proliferation for optimal infection; (2) immune responses, whether pre-existing or developing after vector delivery, may rapidly eliminate transgenic protein expression and prevent future effective intervention. Early gene transfer, in the neonatal or even fetal period, may overcome some or all of these obstacles . Therefore, the experimental approach described herein, might be useful in the development of new approaches to gene therapy in young organisms.
In summary, we describe an eukaryotic dicistronic vector system that is capable of transducing mouse neural precursors in vivo andmaintaining the expression of genes after cell differentiation. Human placental alkaline phosphatase (PLAP) and neomycin phosphotransferase (Neo) used in this study as reporter genes can be replaced by other genes of interest to make these dicistronic vectors a novel tool to trace lineages and assess the function of specific genes in rodent CNS in vivo. Vectors might also be ideally suited to targeting suicide genes to proliferating cells, such as tumor cells, that spread and infiltrate via the CSF [61, 62].
Materials and methods
Vectors, helper cells, titration
Plasmid vectors pEMCV-CBT4, pREV-HW1 and pREV-HW3, shown schematically in Fig 1, have been previously described [22, 23]. NIH-3T3 cells, and the NIH-3T3 based retroviral packaging cell line GP+E-86 , were cultured in Dulbecco's modified Eagle's medium (DMEM, Gibco BRL) with 10% newborn calf serum at 37°C in presence of 5% CO2. MLV vectors were produced by transfection of GP+E-86 cells with pREV-HW3 or pEMCV-CBT4 constructs as previously described . Vectors, produced by GP+E-86, were titrated on NIH-3T3 [15, 16, 22]. The negative control pREV-HW1 was produced using the same procedure as above.
Postnatal day 1–2 mice (OF1 strain) were injected with 1–5 × 104 producer cells in a 2 μl volume in the region of the developing cerebellum as follows. Producer cells harboring the recombinant vector, or control cells (non-transfected helper cells, transduced 3T3 cells or cells transfected with the pRev-HW 1 vector described by López-Lastra et al., (1997) which cannot be packaged, see Fig. 1), were resuspended by trypsinization, washed once in medium and twice in PBS, counted and resuspended in PBS. Cell suspension was pumped from a Hamilton syringe to fill a fine plastic catheter connected to a second Hamilton syringe needle. The second needle was manually pierced to a depth of 1.5 – 2 mm through the cranial cartilage into the region of the developing cerebellum, behind the cerebral hemispheres which were visible through the skull. Then, 2 μl of cell suspension was slowly pumped into the brain using the Hamilton syringe over a 10 second period. The needle was held in place for another 10 seconds and then carefully removed. The young were then replaced with their mothers and maintained with free access to food and water until the time of sacrifice. Animals were killed by anoxia in CO2, decapitated and their brains were carefully removed and fixed by immersion in 4% paraformaldehyde in PBS for 12 – 15 h. Brains were then cryoprotected by immersion in 30% sucrose and frozen by immersion in isopentane over dry ice. Brains were then cut into serial sections of 16 μμm thickness using a Leitz cryomicrotome and recovered on gelatin-coated glass microscope slides. All experiments involving animals were performed in accordance with the French regulations and were approved by the animal experimentation committee of the Ecole Normale Supérieure, Lyon.
For placental alkaline phosphatase (PLAP) histochemical staining, cells were fixed in phosphate-buffered saline (PBS) containing 4% paraformaldehyde. After two washes in PBS, they were incubated at 65°C for 30 min in PBS. Tissue sections were incubated for 1 hour at 65°C. Cells or tissue sections were washed twice with AP buffer (100 mM Tris-HCl pH 9.5, 100 mM NaCl, and 5 mM MgCl2) and incubated for 5 hr in staining solution (0.1 mg/ml 5-bromo-4-chloro-3-indolyl phosphate (BCIP), 1 mg/ml nitroblue tetrazolium salt (NBT), and 1 mM levamisole) at 22°C. Brain regions of histochemically and immunostained cells were identified by extrapolation from a rat brain histological atlas .
Tissue sections were rinsed in PBS then incubated for 30 min in 20 mM ammonium acetate. Sections were washed twice in PBS then incubated for 30 min in a blocking solution of PBS containing 5% BSA, 1% normal goat serum and 0.2% Tween 20 prior to staining with antibodies. This same solution served to dilute all the antibodies. Double-labelling was performed by simultaneous staining with antibodies produced in different species which were then revealed using fluorochrome-conjugated goat antibodies with appropriate species specificity. Thus, PLAP was revealed using a murine monoclonal antibody (diluted 1/200) purchased from DAKO (Glostrup, Denmark). Neo was revealed using an affinity purified rabbit polyclonal antibody (diluted 1/100) generated by immunizing rabbits with peptides VENGRFSGFIDCGRL and MIEQDGLHAGSPAAC conjugated by their carboxy terminus to keyhole limpet haemocyanin. GFAP which labels astrocytes , a population of neural stem cells [27, 58] developing ependymocytes  and radial glia  was detected by a polyclonal rabbit anti-cow GFAP antiserum (diluted 1/200), purchased from DAKO. Neurons were detected using an anti-HU antiserum generously donated by Dr. J. Honnorat and Dr. M-F. Belin (diluted 1/1000). The HU antigen comprises a group of nucleic acid binding proteins located in the nucleus and cytoplasm of post-mitotic neurons . All antibodies have been tested in cell culture and on various control tissues and give appropriate patterns of specific labeling. The neural cell type-specific markers did not label helper/producer cells in vitro. Primary incubations were for 2 h at room temperature or at 4°C overnight. After washing sections 5 × 10 min in PBS, bound antibodies were revealed with FITC-conjugated goat anti-human immunoglobulin antibodies or Cy3-conjugated goat anti-rabbit IgG antibodies and either Cy3- or FITC-conjugated goat anti-mouse IgG antibodies. Anti-immunoglobulin antibodies were all at a final dilution of 1/400 in blocking buffer containing bis-benzimide (1 μg/ml) to stain DNA. Controls included no primary antibodies and non-transduced brain. Slides were washed 3 times in PBS, mounted with moviol and analyzed with a Zeiss Axioplan fluorescence microscope.
The authors wish to thank Christelle Daudé for her expert technical assistance. This work was supported by grants from the ANRS, the MGEN and ARC (number 5466) to J-L Darlix, and the Pontificia Universidad Católica de Chile (DIPUC 2004/06E and 2005/14PI) to M. López-Lastra. E.A. Derrington was supported in part by a fellowship from the ANRS.
- Snyder EY, Taylor RM, Wolfe JH: Neural progenitor cell engraftment corrects lysosomal storage throughout the MPS VII mouse brain. Nature. 1995, 374: 367-370. 10.1038/374367a0.View ArticlePubMedGoogle Scholar
- Martinez-Serrano A, Lundberg C, Horellou P, Fischer W, Bentlage C, Campbell K, McKay RD, Mallet J, Bjorklund A: CNS-derived neural progenitor cells for gene transfer of nerve growth factor to the adult rat brain: complete rescue of axotomized cholinergic neurons after transplantation into the septum. J Neurosci. 1995, 15: 5668-5680.PubMedGoogle Scholar
- Brustle O, Maskos U, McKay RD: Host-guided migration allows targeted introduction of neurons into the embryonic brain. Neuron. 1995, 15: 1275-1285. 10.1016/0896-6273(95)90007-1.View ArticlePubMedGoogle Scholar
- Gage FH, Coates PW, Palmer TD, Kuhn HG, Fisher LJ, Suhonen JO, Peterson DA, Suhr ST, Ray J: Survival and differentiation of adult neuronal progenitor cells transplanted to the adult brain. Proc Natl Acad Sci U S A. 1995, 92: 11879-11883.PubMed CentralView ArticlePubMedGoogle Scholar
- Snyder EY, Deitcher DL, Walsh C, Arnold-Aldea S, Hartwieg EA, Cepko CL: Multipotent neural cell lines can engraft and participate in development of mouse cerebellum. Cell. 1992, 68: 33-51. 10.1016/0092-8674(92)90204-P.View ArticlePubMedGoogle Scholar
- Renfranz PJ, Cunningham MG, McKay RD: Region-specific differentiation of the hippocampal stem cell line HiB5 upon implantation into the developing mammalian brain. Cell. 1991, 66: 713-729. 10.1016/0092-8674(91)90116-G.View ArticlePubMedGoogle Scholar
- Snyder EY, Yoon C, Flax JD, Macklis JD: Multipotent neural precursors can differentiate toward replacement of neurons undergoing targeted apoptotic degeneration in adult mouse neocortex. Proc Natl Acad Sci U S A. 1997, 94: 11663-11668. 10.1073/pnas.94.21.11663.PubMed CentralView ArticlePubMedGoogle Scholar
- Nait-Oumesmar B, Decker L, Lachapelle F, Avellana-Adalid V, Bachelin C, Van Evercooren AB: Progenitor cells of the adult mouse subventricular zone proliferate, migrate and differentiate into oligodendrocytes after demyelination. Eur J Neurosci. 1999, 11: 4357-4366. 10.1046/j.1460-9568.1999.00873.x.View ArticlePubMedGoogle Scholar
- Aboody KS, Brown A, Rainov NG, Bower KA, Liu S, Yang W, Small JE, Herrlinger U, Ourednik V, Black PM, Breakefield XO, Snyder EY: Neural stem cells display extensive tropism for pathology in adult brain: evidence from intracranial gliomas. Proc Natl Acad Sci U S A. 2000, 97: 12846-12851. 10.1073/pnas.97.23.12846.PubMed CentralView ArticlePubMedGoogle Scholar
- Lewis PF, Emerman M: Passage through mitosis is required for oncoretroviruses but not for the human immunodeficiency virus. J Virol. 1994, 68: 510-516.PubMed CentralPubMedGoogle Scholar
- Weber E, Anderson WF, Kasahara N: Recent advances in retrovirus vector-mediated gene therapy: teaching an old vector new tricks. Curr Opin Mol Ther. 2001, 3: 439-453.PubMedGoogle Scholar
- Bao ZZ, Cepko CL: The expression and function of Notch pathway genes in the developing rat eye. J Neurosci. 1997, 17: 1425-1434.PubMedGoogle Scholar
- Gaiano N, Kohtz JD, Turnbull DH, Fishell G: A method for rapid gain-of-function studies in the mouse embryonic nervous system. Nat Neurosci. 1999, 2: 812-819. 10.1038/12186.View ArticlePubMedGoogle Scholar
- Burrows RC, Wancio D, Levitt P, Lillien L: Response diversity and the timing of progenitor cell maturation are regulated by developmental changes in EGFR expression in the cortex. Neuron. 1997, 19: 251-267. 10.1016/S0896-6273(00)80937-X.View ArticlePubMedGoogle Scholar
- Derrington EA, Lopez-Lastra M, Chapel-Fernandez S, Cosset FL, Belin MF, Rudkin BB, Darlix JL: Retroviral vectors for the expression of two genes in human multipotent neural precursors and their differentiated neuronal and glial progeny. Hum Gene Ther. 1999, 10: 1129-1138. 10.1089/10430349950018120.View ArticlePubMedGoogle Scholar
- Franceschini IA, Feigenbaum-Lacombe V, Casanova P, Lopez-Lastra M, Darlix JL, Dalcq MD: Efficient gene transfer in mouse neural precursors with a bicistronic retroviral vector. J Neurosci Res. 2001, 65: 208-219. 10.1002/jnr.1144.View ArticlePubMedGoogle Scholar
- Goldman JE, Zerlin M, Newman S, Zhang L, Gensert J: Fate determination and migration of progenitors in the postnatal mammalian CNS. Dev Neurosci. 1997, 19: 42-48.View ArticlePubMedGoogle Scholar
- Altman J: Postnatal development of the cerebellar cortex in the rat. I. The external germinal layer and the transitional molecular layer. J Comp Neurol. 1972, 145: 353-397. 10.1002/cne.901450305.View ArticlePubMedGoogle Scholar
- Altman J: Postnatal development of the cerebellar cortex in the rat. 3. Maturation of the components of the granular layer. J Comp Neurol. 1972, 145: 465-513. 10.1002/cne.901450403.View ArticlePubMedGoogle Scholar
- Altman J: Postnatal development of the cerebellar cortex in the rat. II. Phases in the maturation of Purkinje cells and of the molecular layer. J Comp Neurol. 1972, 145: 399-463. 10.1002/cne.901450402.View ArticlePubMedGoogle Scholar
- Miale IL, Sidman RL: An autoradiographic analysis of histogenesis in the mouse cerebellum. Exp Neurol. 1961, 4: 277-296. 10.1016/0014-4886(61)90055-3.View ArticlePubMedGoogle Scholar
- Lopez-Lastra M, Gabus C, Darlix JL: Characterization of an internal ribosomal entry segment within the 5' leader of avian reticuloendotheliosis virus type A RNA and development of novel MLV-REV-based retroviral vectors. Hum Gene Ther. 1997, 8: 1855-1865.View ArticlePubMedGoogle Scholar
- Torrent C, Berlioz C, Darlix JL: Stable MLV-VL30 dicistronic retroviral vectors with a VL30 or MoMLV sequence promoting both packaging of genomic RNA and expression of the 3' cistron. Hum Gene Ther. 1996, 7: 603-612.View ArticlePubMedGoogle Scholar
- Dalmau J, Furneaux HM, Cordon-Cardo C, Posner JB: The expression of the Hu (paraneoplastic encephalomyelitis/sensory neuronopathy) antigen in human normal and tumor tissues. Am J Pathol. 1992, 141: 881-886.PubMed CentralPubMedGoogle Scholar
- Rosenblum MK: Paraneoplasia and autoimmunologic injury of the nervous system: the anti-Hu syndrome. Brain Pathol. 1993, 3: 199-212.View ArticlePubMedGoogle Scholar
- Chiasson BJ, Tropepe V, Morshead CM, van der Kooy D: Adult mammalian forebrain ependymal and subependymal cells demonstrate proliferative potential, but only subependymal cells have neural stem cell characteristics. J Neurosci. 1999, 19: 4462-4471.PubMedGoogle Scholar
- Doetsch F, Caille I, Lim DA, Garcia-Verdugo JM, Alvarez-Buylla A: Subventricular zone astrocytes are neural stem cells in the adult mammalian brain. Cell. 1999, 97: 703-716. 10.1016/S0092-8674(00)80783-7.View ArticlePubMedGoogle Scholar
- Garcia AD, Doan NB, Imura T, Bush TG, Sofroniew MV: GFAP-expressing progenitors are the principal source of constitutive neurogenesis in adult mouse forebrain. Nat Neurosci. 2004, 7: 1233-1241. 10.1038/nn1340.View ArticlePubMedGoogle Scholar
- Johansson CB, Momma S, Clarke DL, Risling M, Lendahl U, Frisen J: Identification of a neural stem cell in the adult mammalian central nervous system. Cell. 1999, 96: 25-34. 10.1016/S0092-8674(00)80956-3.View ArticlePubMedGoogle Scholar
- Bruni JE: Ependymal development, proliferation, and functions: a review. Microsc Res Tech. 1998, 41: 2-13. 10.1002/(SICI)1097-0029(19980401)41:1<2::AID-JEMT2>3.0.CO;2-Z.View ArticlePubMedGoogle Scholar
- Frederiksen K, McKay RD: Proliferation and differentiation of rat neuroepithelial precursor cells in vivo. J Neurosci. 1988, 8: 1144-1151.PubMedGoogle Scholar
- Alvarez-Buylla A, Theelen M, Nottebohm F: Proliferation "hot spots" in adult avian ventricular zone reveal radial cell division. Neuron. 1990, 5: 101-109. 10.1016/0896-6273(90)90038-H.View ArticlePubMedGoogle Scholar
- Gray GE, Sanes JR: Lineage of radial glia in the chicken optic tectum. Development. 1992, 114: 271-283.PubMedGoogle Scholar
- Malatesta P, Hartfuss E, Gotz M: Isolation of radial glial cells by fluorescent-activated cell sorting reveals a neuronal lineage. Development. 2000, 127: 5253-5263.PubMedGoogle Scholar
- Hartfuss E, Galli R, Heins N, Gotz M: Characterization of CNS precursor subtypes and radial glia. Dev Biol. 2001, 229: 15-30. 10.1006/dbio.2000.9962.View ArticlePubMedGoogle Scholar
- Kamei Y, Inagaki N, Nishizawa M, Tsutsumi O, Taketani Y, Inagaki M: Visualization of mitotic radial glial lineage cells in the developing rat brain by Cdc2 kinase-phosphorylated vimentin. Glia. 1998, 23: 191-199. 10.1002/(SICI)1098-1136(199807)23:3<191::AID-GLIA2>3.0.CO;2-8.View ArticlePubMedGoogle Scholar
- Halliday AL, Cepko CL: Generation and migration of cells in the developing striatum. Neuron. 1992, 9: 15-26. 10.1016/0896-6273(92)90216-Z.View ArticlePubMedGoogle Scholar
- Kempler G, Freitag B, Berwin B, Nanassy O, Barklis E: Characterization of the Moloney murine leukemia virus stem cell-specific repressor binding site. Virology. 1993, 193: 690-699. 10.1006/viro.1993.1177.View ArticlePubMedGoogle Scholar
- Cornelis S, Bruynooghe Y, Denecker G, Van Huffel S, Tinton S, Beyaert R: Identification and characterization of a novel cell cycle-regulated internal ribosome entry site. Mol Cell. 2000, 5: 597-605. 10.1016/S1097-2765(00)80239-7.View ArticlePubMedGoogle Scholar
- Brasey A, Lopez-Lastra M, Ohlmann T, Beerens N, Berkhout B, Darlix JL, Sonenberg N: The leader of human immunodeficiency virus type 1 genomic RNA harbors an internal ribosome entry segment that is active during the G2/M phase of the cell cycle. J Virol. 2003, 77: 3939-3949. 10.1128/JVI.77.7.3939-3949.2003.PubMed CentralView ArticlePubMedGoogle Scholar
- Pyronnet S, Pradayrol L, Sonenberg N: A cell cycle-dependent internal ribosome entry site. Mol Cell. 2000, 5: 607-616. 10.1016/S1097-2765(00)80240-3.View ArticlePubMedGoogle Scholar
- Tinton SA, Schepens B, Bruynooghe Y, Beyaert R, Cornelis S: Regulation of the cell-cycle-dependent internal ribosome entry site of the PITSLRE protein kinase: roles of Unr (upstream of N-ras) protein and phosphorylated translation initiation factor eIF-2alpha. Biochem J. 2005, 385: 155-163. 10.1042/BJ20040963.PubMed CentralView ArticlePubMedGoogle Scholar
- Lang KJ, Kappel A, Goodall GJ: Hypoxia-inducible factor-1alpha mRNA contains an internal ribosome entry site that allows efficient translation during normoxia and hypoxia. Mol Biol Cell. 2002, 13: 1792-1801. 10.1091/mbc.02-02-0017.PubMed CentralView ArticlePubMedGoogle Scholar
- Stein I, Itin A, Einat P, Skaliter R, Grossman Z, Keshet E: Translation of vascular endothelial growth factor mRNA by internal ribosome entry: implications for translation under hypoxia. Mol Cell Biol. 1998, 18: 3112-3119.PubMed CentralView ArticlePubMedGoogle Scholar
- Kim YK, Jang SK: Continuous heat shock enhances translational initiation directed by internal ribosomal entry site. Biochem Biophys Res Commun. 2002, 297: 224-231. 10.1016/S0006-291X(02)02154-X.View ArticlePubMedGoogle Scholar
- Subkhankulova T, Mitchell SA, Willis AE: Internal ribosome entry segment-mediated initiation of c-Myc protein synthesis following genotoxic stress. Biochem J. 2001, 359: 183-192. 10.1042/0264-6021:3590183.PubMed CentralView ArticlePubMedGoogle Scholar
- Prats AC, Prats H: Translational control of gene expression: role of IRESs and consequences for cell transformation and angiogenesis. Prog Nucleic Acid Res Mol Biol. 2002, 72: 367-413.View ArticlePubMedGoogle Scholar
- Holcik M, Sonenberg N, Korneluk RG: Internal ribosome initiation of translation and the control of cell death. Trends Genet. 2000, 16: 469-473. 10.1016/S0168-9525(00)02106-5.View ArticlePubMedGoogle Scholar
- Holcik M: Translational upregulation of the X-linked inhibitor of apoptosis. Ann N Y Acad Sci. 2003, 1010: 249-258. 10.1196/annals.1299.043.View ArticlePubMedGoogle Scholar
- Van Eden ME, Byrd MP, Sherrill KW, Lloyd RE: Translation of cellular inhibitor of apoptosis protein 1 (c-IAP1) mRNA is IRES mediated and regulated during cell stress. Rna. 2004, 10: 469-481. 10.1261/rna.5156804.PubMed CentralView ArticlePubMedGoogle Scholar
- Henis-Korenblit S, Strumpf NL, Goldstaub D, Kimchi A: A novel form of DAP5 protein accumulates in apoptotic cells as a result of caspase cleavage and internal ribosome entry site-mediated translation. Mol Cell Biol. 2000, 20: 496-506. 10.1128/MCB.20.2.496-506.2000.PubMed CentralView ArticlePubMedGoogle Scholar
- Maier D, Nagel AC, Preiss A: Two isoforms of the Notch antagonist Hairless are produced by differential translation initiation. Proc Natl Acad Sci U S A. 2002, 99: 15480-15485. 10.1073/pnas.242596699.PubMed CentralView ArticlePubMedGoogle Scholar
- Chalandon Y, Jiang X, Hazlewood G, Loutet S, Conneally E, Eaves A, Eaves C: Modulation of p210(BCR-ABL) activity in transduced primary human hematopoietic cells controls lineage programming. Blood. 2002, 99: 3197-3204. 10.1182/blood.V99.9.3197.View ArticlePubMedGoogle Scholar
- Sella O, Gerlitz G, Le SY, Elroy-Stein O: Differentiation-induced internal translation of c-sis mRNA: analysis of the cis elements and their differentiation-linked binding to the hnRNP C protein. Mol Cell Biol. 1999, 19: 5429-5440.PubMed CentralView ArticlePubMedGoogle Scholar
- Ye X, Fong P, Iizuka N, Choate D, Cavener DR: Ultrabithorax and Antennapedia 5' untranslated regions promote developmentally regulated internal translation initiation. Mol Cell Biol. 1997, 17: 1714-1721.PubMed CentralView ArticlePubMedGoogle Scholar
- Bernstein J, Sella O, Le SY, Elroy-Stein O: PDGF2/c-sis mRNA leader contains a differentiation-linked internal ribosomal entry site (D-IRES). J Biol Chem. 1997, 272: 9356-9362. 10.1074/jbc.272.14.9356.View ArticlePubMedGoogle Scholar
- Chauhan AN, Lewis PD: A quantitative study of cell proliferation in ependyma and choroid plexus in the postnatal rat brain. Neuropathol Appl Neurobiol. 1979, 5: 303-309.View ArticlePubMedGoogle Scholar
- Doetsch F, Garcia-Verdugo JM, Alvarez-Buylla A: Cellular composition and three-dimensional organization of the subventricular germinal zone in the adult mammalian brain. J Neurosci. 1997, 17: 5046-5061.PubMedGoogle Scholar
- Levitt P, Rakic P: Immunoperoxidase localization of glial fibrillary acidic protein in radial glial cells and astrocytes of the developing rhesus monkey brain. J Comp Neurol. 1980, 193: 815-840. 10.1002/cne.901930316.View ArticlePubMedGoogle Scholar
- Waddington SN, Kennea NL, Buckley SM, Gregory LG, Themis M, Coutelle C: Fetal and neonatal gene therapy: benefits and pitfalls. Gene Ther. 2004, 11 Suppl 1: S92-7. 10.1038/sj.gt.3302375.View ArticlePubMedGoogle Scholar
- Klatzmann D, Valery CA, Bensimon G, Marro B, Boyer O, Mokhtari K, Diquet B, Salzmann JL, Philippon J: A phase I/II study of herpes simplex virus type 1 thymidine kinase "suicide" gene therapy for recurrent glioblastoma. Study Group on Gene Therapy for Glioblastoma. Hum Gene Ther. 1998, 9: 2595-2604. 10.1089/10430349850019436.View ArticlePubMedGoogle Scholar
- Valery CA, Seilhean D, Boyer O, Marro B, Hauw JJ, Kemeny JL, Marsault C, Philippon J, Klatzmann D: Long-term survival after gene therapy for a recurrent glioblastoma. Neurology. 2002, 58: 1109-1112.View ArticlePubMedGoogle Scholar
- Markowitz D, Goff S, Bank A: A safe packaging line for gene transfer: separating viral genes on two different plasmids. J Virol. 1988, 62: 1120-1124.PubMed CentralPubMedGoogle Scholar
- Paxinos G, Watson C: The rat brain in stereotactic coordinates. Compact third edition. 1997, , Academic Press, San Diego. USA.Google Scholar
- Bignami A, Eng LF, Dahl D, Uyeda CT: Localization of the glial fibrillary acidic protein in astrocytes by immunofluorescence. Brain Res. 1972, 43: 429-435. 10.1016/0006-8993(72)90398-8.View ArticlePubMedGoogle Scholar
- Jang SK, Wimmer E: Cap-independent translation of encephalomyocarditis virus RNA: structural elements of the internal ribosomal entry site and involvement of a cellular 57-kD RNA-binding protein. Genes Dev. 1990, 4: 1560-1572.View ArticlePubMedGoogle Scholar
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