Mutation of conserved aspartates affect maturation of presenilin 1 and presenilin 2 complexes

102

The majority of familial Alzheimer's disease cases have been attributed to mutations in the presenilin 1 (PS1) gene. PS1 is synthesized as an inactive holoprotein that undergoes endoproteolytic processing to generate a functional N-and C-terminal heterodimer (NTF and CTF, respectively). We identified a single residue in PS1, Ser 397 , which regulates the CTF levels in a population of dimer that has a rapid turnover. This residue is part of a highly conserved glycogen synthase kinase-3␤ (GSK-3␤) consensus phosphorylation site within the loop domain of PS1. Site-directed mutagenesis at the Ser 397 position increased levels of PS1 CTF but not NTF or holoprotein. Similar increases in only CTF levels were seen when cells expressing wild type PS1 were treated with lithium chloride, an inhibitor of GSK-3␤. Both wild type and PS1 S397A CTF displayed a biphasic turnover, reflecting rapidly degraded and stable populations. Rapid turnover was delayed for mutant PS1 S397A, causing increased CTF. These data demonstrate that PS1 NTF⅐CTF endoproteolytic fragments are generated in excess, that phosphorylation at Ser 397 by GSK-3␤ regulates the discard of excess CTF, and that the disposal of surplus NTF is mediated by an independent mechanism. Overall, the results indicate that production of active NTF⅐CTF dimer is more complex than limited endoproteolysis of PS1 holoprotein and instead involves additional regulatory events.

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Glycogen Synthase Kinase-3β Regulates Presenilin 1 C-terminal Fragment Levels

Kirschenbaum¹,

Hsu²,

Cordell³

et al. 2001

102

“…4B, lane 5). The PS1 D385A and PS1 D257A mutations and the equivalent mutations in PS2 were shown previously to generate only the intermediary, ϳ180-kDa, immature, non-functional complexes containing holoprotein (25), which do not mature into the higher molecular weight (Ͼ Ͼ250 kDa) functional complex containing PS1-NTF/CTFs (25). The apparently larger amount of APH-1 1 L co-immunoprecipitated with PS1 D385A compared with wild type PS1 (Fig.…”

Section: Fig 2 Western Blot Analysis Of Endogenous and Transfected mentioning

confidence: 99%

APH-1 Interacts with Mature and Immature Forms of Presenilins and Nicastrin and May Play a Role in Maturation of Presenilin·Nicastrin Complexes

Chen

Sanjo

et al. 2003

151

152

APH-1 and PEN-2 genes modulate the function of nicastrin and the presenilins in Caenorhabditis elegans.Preliminary studies in transfected mammalian cells overexpressing tagged APH-1 proteins suggest that this genetic interaction is mediated by a direct physical interaction. Using the APH-1 protein encoded on human chromosome 1 (APH-1 1 L; also known as APH-1a) as an archetype, we report here that endogenous forms of APH-1 are predominantly expressed in intracellular membrane compartments, including the endoplasmic reticulum and cis-Golgi. APH-1 proteins directly interact with immature and mature forms of the presenilins and nicastrin within high molecular weight complexes that display ␥-and ⑀-secretase activity. Indeed APH-1 proteins can bind to the nicastrin ⌬312-369 loss of function mutant, which does not undergo glycosylation maturation and is not trafficking beyond the endoplasmic reticulum. The levels of expression of endogenous APH-1 1 L can be suppressed by overexpression of any other members of the APH-1 family, suggesting that their abundance is coordinately regulated. Finally, although the absence of APH-1 destabilizes the presenilins, in contrast to nicastrin and PEN-2, APH-1 itself is only modestly destabilized in cells lacking functional expression of presenilin 1 or presenilin 2. Taken together, our data suggest that APH-1 proteins, and APH-1 1 in particular, may have a role in the initial assembly and maturation of presenilin⅐nicastrin complexes.Presenilin 1 (PS1) 1 (1), presenilin 2 (PS2) (2), and nicastrin (3) are components of high molecular weight protein complexes that are required for the intramembranous proteolysis of some type 1 transmembrane proteins, including the ␤-amyloid precursor protein (APP) (4), Notch (5-11), and ErbB-4 (12). Genetic screens in Caenorhabditis elegans have identified two additional proteins, APH-1 (13) and PEN-2 (14), in which loss of function mutations phenotypically modulate Notch signaling in a manner similar to that of null mutants in nicastrin and the presenilins. Preliminary studies in transfected cells overexpressing tagged APH-1 proteins suggest that the genetic interaction between APH-1 and the presenilin-dependent cleavage of Notch and APP is mediated by a direct physical interaction between APH-1 and nicastrin or the presenilins (14). We report here that both the APH-1 homologue on human chromosome 1 (termed APH-1 a in Ref. 14, but here referred to as APH-1 1 for clarity to avoid confusion with labeling of alternate splice forms) and the APH-1 homologue on chromosome 15 (previously referred to as APH-1 b ; here termed APH-1 15 ) are widely expressed in multiple tissues and that the APH-1 1 transcript is present as several different alternatively spliced forms (data not shown). We also report that endogenous APH-1 1 directly interacts with both immature and mature forms of the presenilins and nicastrin and that high molecular weight complexes containing these proteins display ␥-and ⑀-secretase activity. Overexpression of any one member of the APH-1 family...

“…The evidence for this view includes the following observations. 1) D257A PSEN1, which could not restore neurotrypsin up-regulation induced by the double deficiency, is incapable of endoproteolytic cleavage and is unable to assemble into a mature ␥-secretase complex (40). 2) The uncleavable ⌬E9 hPSEN1 mutant form, which is still able to associate and form a mature ␥-secretase complex (41), efficiently restored neurotrypsin expression in Psen1/2 dKO cells.…”

Section: Discussionmentioning

confidence: 99%

“…Although these results may suggest a ␥-secretase-independent mechanism to regulate neurotrypsin expression, previous studies revealed that drug inhibitors of ␥-secretase are not necessarily equally efficient in block- ing all ␥-secretase-dependent activities (36 -38). Therefore, to unambiguously test whether ␥-secretase is involved in this process, we took advantage of the catalytically dead D257A hPSEN1 mutant, which is unable to undergo autocatalytic endoproteolytic cleavage and unable to form a mature high molecular weight active ␥-secretase complex (39,40). As expected, unlike reintroduction of wild-type hPSEN1/2, reintroduction of D257A hPSEN1 and wild-type hPSEN2 into Psen1/2 dKO cells showed no detectable PSEN1 CTFs or mature nicastrin (Fig.…”

Section: Tg-prss12 Thymentioning

confidence: 99%

Presenilins Regulate Neurotrypsin Gene Expression and Neurotrypsin-dependent Agrin Cleavage via Cyclic AMP Response Element-binding Protein (CREB) Modulation

Almenar-Queralt

Kim

Benner

et al. 2013

Background: Presenilins regulate multiple cellular pathways by controlling transcription. Results: Presenilins repress neurotrypsin expression by preventing CREB recruitment to its promoter. Conclusion: A new strategy utilized by presenilins to regulate CREB signaling relies on preventing CREB recruitment to gene promoters. Significance: Learning how presenilins control CREB signaling will help understanding their physiological/pathological roles.