Two Peptides Derived from the Nerve Growth Factor Precursor Are Biologically Active

Abstract

This report provides evidence that the proregion of the NGF precursor protein contains two novel bioactive peptides. The presence of pairs of basic amino acid (aa) residues in the NGF proregion suggests that two or three peptides other than NGF may be generated by proteolytic cleavage. Synthetic peptides of 29 aa (LIP1) and 38aa (LIP2) corresponding to the sequences −71 to −43 and −40 to −3 of the proNGF, respectively, were used in this study. ELISA specific for these two peptides revealed their presence in the rat intestine. LIP1 was localized by immunohistochemistry in endocrine cells of the intestinal epithelium, and LIP2 was immunoprecipitated from an intestinal extract. We also provide evidence for the presence of specific receptors for LIP2 in several cell lines. Scatchard analysis indicated the presence of a low affinity binding site with a Kd of ∼10−7 M and a high affinity binding site of 10−9 M. Cross-linking studies revealed receptor forms of about 140 kD and 93 kD in a prostatic adenocarcinoma cell line.LIP1 and LIP2 induced rapid F-actin redistribution in PC12 cells within 2 min of incubation, which suggests a role of LIP1 and LIP2 in the process of neurite outgrowth. Furthermore, both propeptides induced rapid tyrosine phosphorylation of the Trk protein in both prostatic adenocarcinoma cells and PC12 cells, thus implicating trk in their mechanism of action. These results support our hypothesis that two peptides within the NGF precursor protein are biologically active.

Connective Tissue Growth Factor CCN2 Interacts with and Activates the Tyrosine Kinase Receptor TrkA

“…These are proteolytically cleaved in the middle to release the biologically active 12-to 14-kD C-terminal mature forms (49). It is proposed that the N-terminal domain allows for correct protein folding and secretion (49,50), and it may also possess some biologic activities (51). However, secreted unprocessed immature or proneurotrophins are often present in abundance (52).…”

Characterization of nerve growth factor precursor protein expression by human prostate stromal cells: A role in selective neurotrophin stimulation of prostate epithelial cell growth

“…These observations suggest that NGF may be the predominant neurotrophic growth factor for prostate growth. Since the NGF propeptides, L38 and N4, have been observed to induce early NGF responses, including Trk phosphorylation [18], and these propeptide domains occur within prepro-NGF, we investigated whether the L38 and N4 peptides could stimulate proliferation of TSU-pr1 cells. It is clear that these NGF propeptides, at biologically active concentrations [18], did not promote colony formation by TSU-pr1 cells.…”

“…Since the NGF propeptides, L38 and N4, have been observed to induce early NGF responses, including Trk phosphorylation [18], and these propeptide domains occur within prepro-NGF, we investigated whether the L38 and N4 peptides could stimulate proliferation of TSU-pr1 cells. It is clear that these NGF propeptides, at biologically active concentrations [18], did not promote colony formation by TSU-pr1 cells. Thus, it is likely that stimulation of prostate epithelial cell proliferation is a late response which requires the presence of the NGF␤ moiety.…”

“…NGF can be translated as long (35-kDa) and short (27-kDa) precursors (prepro-NGF), which potentially can be proteolytically cleaved at dibasic sites (denoted by underside arrows) to generate proforms of 22-kDa, 18-kDa, and mature 13-kDa NGF␤. The locations of three putative glycosylation sites (denoted by arrowheads) are also shown (adapted from Ullrich et al [3], Dicou [5], and Dicou et al [18]). …”

“…Hence, the biological activity of NGF must be exclusively mediated via the Trk receptor in these prostate tumor cell lines. Moreover, chemically synthesized peptides corresponding to the N4 and L38 domains of the NGF precursor also exhibit biological activity in that they stimulate phosphorylation of the Trk receptor, as well as F-actin redistribution, an early event in neurite formation [18]. This raises the possibility that an NGF precursor protein containing the N4, L38, and NGF␤ moieties could exhibit multiple biological activities on a tumor cell expressing the Trk receptor.…”