Progranulin-derived granulin E and lysosome membrane protein CD68 interact to reciprocally regulate their protein homeostasis.

Progranulin (PGRN) is a glycoprotein implicated in several neurodegenerative diseases. It is highly expressed in microglia and macrophages and can be secreted or delivered to the lysosome compartment. PGRN comprises 7.5 granulin repeats and is processed into individual granulin peptides within the lysosome, but the functions of these peptides are largely unknown. Here, we identify CD68, a lysosome membrane protein mainly expressed in hematopoietic cells, as a binding partner of PGRN and PGRN-derived granulin E. Deletion analysis of CD68 showed that this interaction is mediated by the mucin-proline-rich domain of CD68. While CD68 deficiency does not affect the lysosomal localization of PGRN, it results in a specific decrease in the levels of granulin E but no other granulin peptides. On the other hand, the deficiency of PGRN, and its derivative granulin peptides, leads to a significant shift in the molecular weight of CD68, without altering CD68 localization within the cell. Our results support that granulin E and CD68 reciprocally regulate each other's protein homeostasis.

Differential regulation of progranulin derived granulin peptides.

BACKGROUND: Haploinsufficiency of progranulin (PGRN) is a leading cause of frontotemporal lobar degeneration (FTLD). PGRN is comprised of 7.5 granulin repeats and is processed into individual granulin peptides in the lysosome. However, very little is known about the levels and regulations of individual granulin peptides due to the lack of specific antibodies. RESULTS: Here we report the generation and characterization of antibodies specific to each granulin peptide. We found that the levels of granulins C, E and F are regulated differently  compared to granulins A and B in various tissues. The levels of PGRN and granulin peptides vary in different brain regions and the ratio between granulins and PGRN is highest in the cortical region in the adult male mouse brain. Granulin-A is localized in the lysosome in both neurons and microglia and its levels in microglia increase under pathological conditions. Interestingly,  the levels of granulin A in microglia change correspondingly with PGRN in response to stroke but not demyelination. Furthermore, deficiency of lysosomal proteases and the PGRN binding partner prosaposin leads to alterations in the ratios between individual granulin peptides. Granulins B, C and E are heavily glycosylated and the glycosylation patterns can be regulated. CONCLUSION: Our results support that the levels of individual granulin peptides are differentially regulated under physiological and pathological conditions and provide novel insights into how granulin peptides function in the lysosome.

A multifaceted role of progranulin in regulating amyloid-beta dynamics and responses.

Haploinsufficiency of progranulin (PGRN) is a leading cause of frontotemporal lobar degeneration (FTLD). PGRN polymorphisms are associated with Alzheimer's disease. PGRN is highly expressed in the microglia near Aß plaques and influences plaque dynamics and microglial activation. However, the detailed mechanisms remain elusive. Here we report that PGRN deficiency reduces human APP and Aß levels in the young male but not female mice. PGRN-deficient microglia exhibit increased expression of markers associated with microglial activation, including CD68, galectin-3, TREM2, and GPNMB, specifically near Aß plaques. In addition, PGRN loss leads to up-regulation of lysosome proteins and an increase in the nuclear localization of TFE3, a transcription factor involved in lysosome biogenesis. Cultured PGRN-deficient microglia show enhanced nuclear translocation of TFE3 and inflammation in response to Aß fibril treatment. Taken together, our data revealed a sex- and age-dependent effect of PGRN on APP metabolism and a role of PGRN in regulating lysosomal activities and inflammation in plaque-associated microglia.