Genetic mutations inGranulin (GRN) that result in reduced levels of its encodedprotein,progranulin (PGRN), have been implicated in several distinctneurological disorders,dependingon thedegree ofPGRN reduction. More specifically, haploinsufficiency resulting from heterozygous GRN mutations has been identified to be causal for a subset of patients with frontotemporal lobar degeneration (FTLD), an adult-onset neurodegenerative disease. Furthermore, homozygous loss-of-function GRN mutations result in neuronal ceroid lipofuscinosis (NCL), and single nucleotide variants that decrease plasma and brain PGRN levels are risk factors for Alzheimer?s Disease (AD). The clear association between reduced levels of PGRN and neurological disorders highlights the importance of adequate PGRN dosage in normal nervous system function. The overarching goal of this project is to better understand the precise cellular and molecular mechanisms that are involved in the regulation of PGRN. In order to reach this goal, we began by identifying factors that could potentially modulate phenotypes associated with PGRN haploinsufficiency. The preliminary data presented in this application clearly show that Nemo-like kinase (NLK), an evolutionarily conserved serine/threoninekinase,isinvolvedintheregulationofPGRNlevelsandcanmodulatephenotypesassociated with PGRN reduction in vivo through microglia. To investigatethis ideafurther, we propose thefollowing three specific aims.
In Specific Aim 1, we will determine whether altering Nlk levels specifically in microglia can modulate FTLD-related phenotypes in vivo using mouse genetics. Specifically, (1) we will first test whether constitutive loss of Nlk in microglia is able to induce FTLD-related neuropathological and behavioral phenotypes. (2) Conversely, we will examine if constitutive overexpression of Nlk in microglia can prevent or ameliorate these same phenotypes. We will focus on neuropathological changes and behavioral deficits that havebeenpreviouslyascribedtoPGRNreductioninFTLD-PGRNpatientsandGrnknockoutmice.
In Specific Aim2, wewillelucidatethemolecularmechanismthroughwhichNlkregulatesPgrnlevelsinmicroglia.Wewill (1) employ mouse genetics and isogenic human induced pluripotent stem cell-derived microglia to determine the receptor involved in Nlk-mediated regulation of Pgrn endocytosis and (2) utilize unbiased proteomics approaches to identify direct molecular targets of Nlk that function in this regulation.
In Specific Aim 3, we will determine whether GRN-associated neuropathology can be suppressed or reversed by increasing Nlk expression in adulthood. To do this, we will temporally induce the overexpression of Nlk after disease onset and test the efficacy in ameliorating or reversing pathological and behavioral deficits. We believe that the knowledge gained from the studies proposed in this application will advance our basic understanding of the cellular and mechanism underlying PGRN regulation and will suggest new therapeutic interventions aimed at reducingtheburdenofFTLDandotherneurologicaldisordersassociatedwithPGRNreductionsuchasAD.
The burden of Alzheimer?s disease (AD) and Alzheimer?s Disease-Related Dementias (ADRD) is enormous, yetnoeffectivetherapeuticsexistforamajorityofdiseasesymptoms.Thestudiesoutlinedinthisproposalwill allow us to determine the cellular and molecular mechanisms underlying microglial regulation of Progranulin levels and associated neuropathological phenotypes, and therefore may lead to the development of improved treatmentsforneurodegenerativediseasesresultingfromProgranulinreductioninthefuture.
