Distribution, levels and phosphorylation of Raf‐1 in Alzheimer's disease

Abstract

Extracellular signal-regulated kinase (ERK), a member of the mitogen-activated protein kinase pathway, has been increasingly implicated in the pathogenesis of Alzheimer's disease due to its critical role in brain function. While we previously demonstrated that ERK is activated in Alzheimer's disease, the upstream cascade leading to its activation had not been fully examined. In this study, we focused on Raf-1, one of the physiological activators of the ERK pathway. Raf-1 is activated by phosphorylation at Ser338 and Tyr340/341 and inhibited by phosphorylation at Ser259. Interestingly, phosphorylation at all three sites on Raf-1 was increased as evidenced by both immunocytochemistry and immunoblot analysis in Alzheimer's disease brains compared to agematched controls. Both phospho-Raf-1 (Ser259) and phospho-Raf-1 (Ser338) were localized to intracytoplasmic granular structures, whereas phospho-Raf-1 (Tyr340/341) was localized to neurofibrillary tangles and granules in pyramidal neurons in Alzheimer's disease hippocampus. There is extensive overlap between phospho-Raf-1 (Ser338) and phospho-Mek1/2, the downstream effector of Raf-1, suggestive of a mechanistic link. Additionally, increased levels of Raf-1 are associated with Ras and MEK1 in Alzheimer's disease as evidenced by its coimmunoprecipitation with Ras and Mek1, respectively. Based on these findings, we speculate that Raf-1 is activated to effectively mediate Ras-dependent signals in Alzheimer's disease.

A Genome-Scale RNA–Interference Screen Identifies RRAS Signaling as a Pathologic Feature of Huntington's Disease

“…This is in agreement with a previous report [45] of enhanced ERK activity in two mutant Htt cell lines, although in that study over-expression of a constitutively active mutant of MEK was protective against caspase induction. We examined the phosphorylation status of RAF1 at S338 in these cellular HD models to determine if signaling is dysregulated, as has been observed previously in Alzheimers disease (AD) patient brains [46] and a mouse model of AD [47]. The ratio of phosphorylated S338 RAF1 to total is higher in ST Hdh Q111/Q111 cells than wild-type, and knock-down of RRAS restores the ratio (Figure 5A).…”

“…The alterations in RAF1 signaling observed in response to mutant Htt expression are consistent with a previous report [45] of enhanced ERK activity in two mutant Htt cell lines, although in that study over-expression of a constitutively active mutant of MEK was protective against caspase induction. Elevated RAF1 S338 phosphorylation has been reported previously in Alzheimer's disease (AD) patient brains [46] and a mouse model of AD [47]. GW5074, an in vitro RAF1 kinase inhibitor, has been shown to have neuroprotective effects in an AD model [47] and against a variety of toxic insults [60].…”

Concerted Perturbation Observed in a Hub Network in Alzheimer’s Disease

“…Abnormal phosphorylation of CDK5 has also been implicated in the formation of amyloid and NFT [50]. Increased levels of RAF1 can effectively mediate Ras-dependent signals and play a critical role in the aberrant activation of the MEK/ERK pathway in AD [51]. BCR [52], [53] and CSK [54], [55], [56] variants have also been associated with AD.…”

Mutation in E1, the Ubiquitin Activating Enzyme, Reduces Drosophila Lifespan and Results in Motor Impairment

“…Therefore, another possibility is that upon aging, specific signaling pathways are dysregulated and contribute to reduced lifespan. In fact, examination of the brains of AD patients found evidence of increased Ras signaling [71]–[75]. Also, expressing activated Ras in neurons causes AD-type phenotypes in neurons in culture [56].…”