| Progranulin (PC-cell derived growth factor, acrogranin, granulin-epithelin precursor) is a secreted glycoprotein with growth promoting activities for diverse cell types and is the precursor for the 6 kDa granulin peptides. These peptides were initially identified in inflammatory cells i.e. neutrophils; and this led to the hypothesis that the grn(granulin ) gene and its products may be regulated during, and function in, wound repair. In response to tissue injury, the grn gene is induced in endothelial cells and fibroblasts. Expression was observed in epithelial and inflammatory cells with higher expression in the inflammatory cells in the early phases of wound healing. This is the first evidence that demonstrates grn gene regulation in an adult physiological state. Given the high expression of the grn gene in hematopoietic cells, particularly the cells of the myeloid lineage, and that these cells are a source of granulin peptides, it was important to understand the signals that regulate progranulin transcript levels in these cells. For this investigation, two myelogenous cell lines, U937 (human histiocytic lymphoma) and HL-60 (human promyelocytic), which are models for monocytic and granulocytic maturation were chosen. Both these lineage related cell lines displayed differential regulation of the progranulin mRNA. The two cell lines were responsive to all three of the differentiation agents, investigated; all-trans retinoic acid (ATRA), dimethylsulphoxide (DMSO) and phorbol-12-myristate-13-acetate (PMA) with the exception of PMA-treated HL-60 cells which exhibit no change in the progranulin transcript levels. As differentiation occurs, the levels of progranulin transcript are up-regulated. However, progranulin does not stimulate the differentiation of these cells as assessed by the expression of the surface marker, CD11b which suggests that the increased levels of progranulin transcript is likely to be a characteristic of matured cells (inflammatory cells). All three differentiation agents increased the half life of the progranulin mRNA in U937 cells, implying that mRNA stability plays an important role in regulating progranulin mRNA levels in these cells. This differs in HL-60 cells, where ATRA and DMSO reduced the transcript's half life. Examination of some of the signaling pathways involved in the regulation of the progranulin transcript level in these cells revealed a highly complex and variable mode of regulation between the two cell lines. The elevation of progranulin mRNA levels in HL-60 cells exposed to ATRA required de novo protein synthesis and was dependent on NF-κB signaling pathways. In contrast, the early increase of progranulin mRNA levels in ATRA-treated U937 cells was independent of protein synthesis. Moreover, it was not impaired by chemical blockade of NF-κB activation. Inhibition of NF-κB pathway did however attenuate progranulin transcript levels induced by PMA in U937 cells. HL-60 cells, but not U937 cells exposed to DMSO displayed a requirement for a protein kinase C (PKC) activation signal to up-regulate progranulin transcript levels. Interestingly, in the absence of serum, DMSO acted as a suppressor of progranulin mRNA levels in HL-60 cells. IL-4 down-regulated basal progranulin transcript levels and it completely abolished the DMSO stimulus in U937 cells. This is the first example of negative regulation of progranulin mRNA levels by a physiological mediator. Other cytokines that activate JAK-STAT signaling pathways also exhibited differential regulation of progranulin mRNA in these cells. GM-CSF displayed minor inhibition of basal mRNA levels in both cell lines while G-CSF had no effects at all. In conclusion, in these two hematopoietic cell lines, increased mRNA levels are associated with a more matured phenotype and is uncoupled from differentiation. ATRA, DMSO and PMA regulate progranulin transcript levels in a cell specific manner. These agents appear to act via NF-κB, and PKC, amongst other pathways but utilize these mechanisms in distinct fashion. The overall level of progranulin mRNA in these cells is likely to be a net result of positive and negative regulatory signals. |
