Evidence for the progression through S-phase in the ectopic cell cycle re-entry of neurons in Alzheimer disease

Abstract

Aberrant neuronal re-entry into the cell cycle is emerging as a potential pathological mechanism in Alzheimer disease (AD). However, while cyclins, cyclin dependent kinases (CDKs), and other mitotic factors are ectopically expressed in neurons, many of these proteins are also involved in other pathological and physiological processes, generating continued debate on whether such markers are truly indicative of a bona fide cell cycle process. To address this issue, here we analyzed one of the minichromosome maintenance (Mcm) proteins that plays a role in DNA replication and becomes phosphorylated by the S-phase promoting CDKs and Cdc7 during DNA synthesis. We found phosphorylated Mcm2 (pMcm2) markedly associated with neurofibrillary tangles, neuropil threads, and dystrophic neurites in AD but not in aged-matched controls. These data not only provide further evidence for cell cycle aberrations in AD, but the cytoplasmic, rather than nuclear, localization of pMcm2 suggests an abnormal cellular distribution of this important replication factor in AD that may explain resultant cell cycle stasis and consequent neuronal degeneration.

Keywords: Alzheimer disease; DNA replication; cell cycle; minichromosome maintenance protein; neurodegeneration.

Figure 1.

In an 87 year old AD case, hippocampal tissue sections demonstrate significant localization of pMcm2 protein in NFT, dystrophic neurites, and neuropil threads (B). In another AD case, in the CA3 region, in addition to pathological structures, a few pyramidal neuron nuclei (arrows) have significant pMcm2 accumulation (D). Most control cases, representative case age 61 years, demonstrate no neuronal staining for pMcm2 protein (A), while a few older control cases demonstrate significant nuclear immunolocalization in the pyramidal neurons (control case age 74 years, C) Scale bar= 50 μm.

Figure 2.

In another AD case, age 63, adjacent hippocampal tissue sections demonstrate many of the AD-related pathological structures (arrows) containing pMcm2 (A) are also positive for hyper-phosphorylated tau (B) in the CA1 region. Lower magnification of adjacent sections of the subiculum shows the large number of NFT and plaques recognized by pMcm2 (C) and AT8 (D). * denotes landmark vessel. Scale bars= 50 μm (A,B), 100 μm (C,D).

Figure 3.

Adsorption of pMcm2 antibody confirms specificity to corresponding pMcm2 antigen. (A) AD hippocampal tissue stained with pMcm2 antibody. (B) Adjacent section treated with pMcm2 antibody absorbed with non-phosphorylated Mcm peptide demonstrates similar staining. (C) Adjacent section treated with pMcm2 antibody absorbed with phosphorylated Mcm2 peptide demonstrates complete absorption. * denotes landmark vessel.

Figure 4.

Pretreatment with alkaline phosphatase to remove phosphate groups, results in elimination of pMcm2 reactivity (B) compared to an untreated adjacent serial section of an AD case (A). * denotes landmark vessel. Scale bar = 50 μm.