Hypothalamic expression of a novel gene product, VGF: immunocytochemical analysis

Pol, AN van den; Decavel, C; Levi, Andrea; Paterson, Bruce M.

doi:10.1523/jneurosci.09-12-04122.1989

Cited by 88 publications

(69 citation statements)

References 38 publications

(47 reference statements)

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“…vgf is developmentally regulated: its expression is detected in embryonic central and peripheral nervous tissues, peaks in the early postnatal critical period of nervous system development, and is maximal during synaptogenesis of the visual cortex (30,41). In adult animals, vgf is expressed in subsets of brain neurons, in sensory and sympathetic neurons, and in endocrine cells from the adrenal medulla, adenohypophysis, and gut (14,49). The VGF protein is stored in secretory vesicles, released in response to stimuli causing membrane depolarization, and proteolytically processed in smaller peptides that may play a role in intercellular communication (39,47).…”

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Interplay of the E Box, the Cyclic AMP Response Element, and HTF4/HEB in Transcriptional Regulation of the Neurospecific, Neurotrophin-InduciblevgfGene†

Rocco

Pennuto

Illi

et al. 1997

Molecular and Cellular Biology

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vgf is a neurotrophin response-specific, developmentally regulated gene that codes for a neurosecretory polypeptide. Its transcription in neuronal cells is selectively activated by the neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor, and neurotrophin 3, which induce survival and differentiation, and not by epidermal growth factor. We studied a short region of the rat vgf promoter which is essential for its regulated expression. A cyclic AMP response element (CRE) within this region is necessary for NGF induction of vgf transcription. Two sites upstream of CRE, an E box and a CCAAT sequence, bind nuclear protein complexes and are involved in transcriptional control. The E box has a dual role. It acts as an inhibitor in NIH 3T3 fibroblasts, together with a second E box located downstream, and as a stimulator in the NGF-responsive cell line PC12. By expression screening, we have isolated the cDNA for a basic helix-loop-helix transcription factor, a homolog of the HTF4/HEB E protein, that specifically binds the vgf promoter E box. The E protein was present in various cell lines, including PC12 cells, and was a component of a multiprotein nuclear complex that binds the promoter in vitro. The E box and CRE cooperate in binding to this complex, which may be an important determinant for neural cell-specific expression.

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Interplay of the E Box, the Cyclic AMP Response Element, and HTF4/HEB in Transcriptional Regulation of the Neurospecific, Neurotrophin-InduciblevgfGene†

Rocco

Pennuto

Illi

et al. 1997

Molecular and Cellular Biology

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“…The constructs described here should allow the appropriate targeting of antibody expression. In particular, the VGF8a gene (23) encodes a secreted protein that is highly expressed in many areas of the nervous system where SP is also found (34,35 3) and the other chain under the control of a tissue-specific promoter. A more localized expression of the antibodies might be achieved by grafting embryonic cells infected with a recombinant retrovirus harboring the sequences that code for this antibody.…”

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confidence: 99%

Neuroantibodies: molecular cloning of a monoclonal antibody against substance P for expression in the central nervous system.

Piccioli

Ruberti

Biocca

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

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We present a strategy to study functional and/or developmental processes occurring in the nervous system, as well as in other systems, of mice. This strategy is based on the local expression of specific monoclonal antibodies (mAbs) by cells of the nervous system. As an application of this strategy, we report the cloning of the anti-substance P rat mAb NC1/34HL. Functional substance P-binding antibodies were reconstituted from the cloned variable domains by using vectors for expression in myeloma cells. With these and other vectors a general system for the cloning and expression of mAbs under a series of promoters (of the rat VGF8a gene, the neurofilament light-chain gene, and the methallothionein gene) has been created. The activity of these plasmids was confirmed by expressing the recombinant NC1/34HL mAb in GH3 pituitary cells, PC12 pheochromocytoma cells, and COS cells. DNA from the described constructs can be used to target the expression of the NC1/34HL mAb to the central nervous system of transgenic mice. This procedure will allow us to perturb substance P activity in a controlled way in order to dissect its multiple roles. afferents is responsible for vasodilatation and plasma extravasation in neurogenic inflammation. SP is also present in central nervous system (CNS) areas, such as the substantia nigra and hypothalamus, where its function is unknown, and in peripheral neuronal structures, where it helps regulate smooth muscle motility. Evidence suggesting roles for SP as a growth factor (7) or as a messenger between the nervous and the immune systems (8) has also been presented. We report here the construction ofgeneral vectors for the expression of any mAb of interest in nonlymphoid mammalian cells, particularly in cells of the nervous system. These vectors have been used to clone the variable regions of the NC1/ 34HL mAbt (9), which recognizes the COOH-terminal part of SP. The NC1/34HL mAb inhibits the interaction of SP with its receptors in vitro as well as in vivo. Several different stable cell lines expressing the cDNA sequence encoding the NC1/ 34HL mAb were derived, and the antigen specificity of the antibody secreted by these cells was confirmed.

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“…Although VGF was initially characterized as a gene product induced in PC12 cells by nerve growth factor (NGF) (58), this peptide is strongly expressed by both the ventrolateral and dorsomedial SCN, but only weakly in the neighboring magnocellular paraventricular and supraoptic nuclei (57)(58)(59). This also supports the "SCN-like," rather than general hypothalamic, character of the SCN 2.2 cell line.…”

Section: Discussionmentioning

confidence: 76%

“…Cross-reactivity with closely related DARPP-32 was not detected in either type of extract (55,56). Additionally, VGF, which is strongly expressed in the SCN, but only weakly in the neighboring magnocellular paraventricular (PVN) and supraoptic nuclei (SON) of the hypothalamus was also used as a SCN marker peptide (57)(58)(59). Both SCN tissue and SCN 2.2 cells expressed VGF-immunoreactive bands at 90 kDa (Fig.…”

Section: Resultsmentioning

confidence: 96%

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Immortalized Suprachiasmatic Nucleus Cells Express Components of Multiple Circadian Regulatory Pathways

Hurst

Earnest

Gillette

2002

Biochemical and Biophysical Research Communications

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We undertook an extensive antigenic characterization of the SCN 2.2 cell line in order to further evaluate whether the line expresses components of circadian regulatory pathways common to the hypothalamic suprachiasmatic nucleus (SCN), the central circadian clock in mammals. We found that differentiated SCN 2.2 cultures expressed a broad range of putative clock genes, as well as components of daytime, nighttime, and crepuscular circadian regulatory pathways found within the SCN in vivo. The line also exhibits several antigens that are highly expressed in a circadian pattern and/or differentially localized in the SCN relative to other hypothalamic regions. Expression of a broad complement of circadian regulatory proteins and putative clock genes further support growing evidence in recent reports that the SCN 2.2 cell line is an appropriate model for investigating the regulation of central mammalian pacemaker. © 2002 Elsevier Science (USA) Key Words: period; cryptochrome; casein kinase; PACAP; PKA; PKC; PKG; NOS; I1; VGF.The central biological pacemaker in mammals lies in the hypothalamic suprachiasmatic nucleus (SCN). The SCN drives circadian behavior, expresses rhythmic gene expression and integrates external stimuli in order to synchronize molecular timekeeping mechanisms with changing environmental conditions. Receptivity to phase shifting stimuli is gated by the circadian state of the central pacemaker. This gating behavior is most clearly demonstrated by the sensitivity of the SCN to light during the night phase, but not during the day phase. All photic, social and hormonal influences on the circadian clock are integrated at the cellular level through multiple signaling pathways within the SCN.The complexity of interacting signaling pathways involved in regulation of the SCN has encouraged researchers to seek cell culture systems to complement in vivo analysis of the mammalian circadian clock. Whether a cell line exhibits central or peripheral circadian clock characteristics complicates selection of an appropriate model system. To provide useful applications for central mammalian pacemaker research, cell line models must exhibit persistent self-sustained oscillations in circadian gene products, restore animal circadian rhythms in SCN-lesioned hosts and exhibit time-dependent responses to stimuli through regulatory pathways that characterize SCN function. Rhythmic gene expression has been demonstrated in NIH/ 3T3 fibroblasts, Rat-1 fibroblasts and spontaneously immortalized embryonic mouse fibroblasts after synchronizing signals (1-5). However, there is no published evidence that these lines express spontaneous, self-sustained rhythms or rescue rhythms in SCNlesioned animals (5, 6). In contrast, another cell line derived from fetal rat SCN, the SCN 2.2 cell line, exhibits endogenous spontaneous circadian rhythmicity in vitro (5, 7). We have focused on the SCN 2.2 line to further assess its usefulness as a model system for studying the mammalian central circadian pacemaker.The SCN 2.2 cell line is a pluripoten...

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Hypothalamic expression of a novel gene product, VGF: immunocytochemical analysis

Abstract: We thank Ms. A. Schneider for technical assistance, and Drs. F. Gage, E. Johnson and E. Shooter for nerve growth factor receptor antibody and suggestions related to it, and Drs. U. di Porzio, R. Vogt, and M. Bennett for useful discussion.

Cited by 88 publications

References 38 publications

Interplay of the E Box, the Cyclic AMP Response Element, and HTF4/HEB in Transcriptional Regulation of the Neurospecific, Neurotrophin-InduciblevgfGene†

Interplay of the E Box, the Cyclic AMP Response Element, and HTF4/HEB in Transcriptional Regulation of the Neurospecific, Neurotrophin-InduciblevgfGene†

Neuroantibodies: molecular cloning of a monoclonal antibody against substance P for expression in the central nervous system.

Immortalized Suprachiasmatic Nucleus Cells Express Components of Multiple Circadian Regulatory Pathways

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