Distinct anterograde trafficking pathways of BACE1 and amyloid precursor protein from the TGN and the regulation of amyloid-β production

Extended residency of BACE1 at early endosomes leads to an increase in Aβ production.

“an extended residency time of BACE1 at the early endosomes also leads to an increase in Aβ production (Toh et al., 2018).”

• cites Toh et al., 2018 sourceSupported

γ-secretase cleavage of C99 releases pathogenic amyloidogenic Aβ associated with Alzheimer's disease.

“The subsequent cleavage of C99 by γ-secretase leads to the release of the pathogenic amyloidogenic beta-amyloid peptide (Aβ) associated with Alzheimer’s disease (Qi-Takahara et al., 2005; Takami et al., 2009; Tomita, 2014).”

• cites Qi-Takahara et al., 2005 sourceSupported
• cites Takami et al., 2009 sourceSupported
• cites Tomita, 2014 sourceNot checked — no full text of the source

Highest BACE1 activity is detected in acidic compartments along the secretory and endocytic pathways including endosomes and trans-Golgi.

“which is consistent with highest BACE1 activity being detected in acidic compartments along the secretory and endocytic pathways including the endosomes and trans-Golgi (Bennett et al., 2000; Huse et al., 2000, 2002; Toh et al., 2017, 2018).”

• cites Bennett et al., 2000 sourceNot checked — no full text of the source
• cites Huse et al., 2000 sourceNot checked — no full text of the source
• cites Toh et al., 2017 sourcePartly supported

  The Introduction identifies early endosomes as a major site of APP processing by BACE1, and the Discussion says the slightly acidic trans-Golgi network provides favourable conditions for BACE1 cleavage. The paper supports activity in those compartments, but does not establish that BACE1 activity is highest there. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Post-Golgi export of BACE1 is independent of the Arl5b/AP-4 sorting pathway.

“As the post-Golgi export of BACE1 is independent of the Arl5b/AP-4 sorting pathway”

• cites Toh et al., 2017 sourceSupported

The presence of BACE1-SBP-GFP in late endosomes/lysosomes suggests turnover of BACE1 in the lysosomes.

“suggesting turnover of BACE1 in the lysosomes (Ye and Cai, 2014; Ye et al., 2017; Feng et al., 2017).”

• cites Ye and Cai, 2014 sourceSupported
• cites Ye et al., 2017Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Feng et al., 2017 sourceSupported

Depletion of Arl5b or AP-4 leads to accumulation of APP, but not BACE1, in the TGN.

“The depletion of either Arl5b or AP-4 leads to the accumulation of APP, but not BACE1, in the TGN.”

• cites Toh et al., 2017 sourceSupported

Newly synthesized APP traffics directly from the TGN to early endosomes.

“We have previously shown that newly synthesized APP traffics directly from the TGN to the early endosomes (Toh et al., 2017)”

• cites Toh et al., 2017 sourceSupported

APP TGN-to-early endosome transport is regulated by AP-4 and Arl5b.

“a transport step regulated by the adaptor AP-4 and the small G protein Arl5b (Burgos et al., 2010; Toh et al., 2017).”

• cites Burgos et al., 2010 sourceSupported
• cites Toh et al., 2017 sourceSupported

Arl5b/AP-4 depletion increases APP processing by endogenous BACE1.

“Arl5b/AP-4 depletion also resulted in an increase in APP processing by endogenous BACE1”

• cites Toh et al., 2017 sourceSupported

BACE1 cleavage of APP is the rate-limiting step along the amyloidogenic processing pathway.

“BACE1 cleavage of APP is the rate-limiting step along the amyloidogenic processing pathway (Thinakaran and Koo, 2008; Zhang et al., 2011)”

• cites Thinakaran and Koo, 2008 sourceNot checked — no full text of the source
• cites Zhang et al., 2011 sourcePartly supported

  In Background > APP Processing, the paper identifies BACE1 cleavage as the first step in Aβ generation and says BACE1 activity is thought to be rate-limiting. The citing sentence presents that qualified assessment as an established fact. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

The TGN is a major site for Aβ production along the secretory pathway.

“We and others have shown that the TGN is a major site for Aβ production along the secretory pathway (Burgos et al., 2010; Toh et al., 2017).”

• cites Burgos et al., 2010 sourcePartly supported

  The Discussion (page not available in our copy) says the authors’ observations support Aβ production in the Golgi complex and/or late secretory pathway: reduced endosomal localization of APP was accompanied by increased Aβ secretion. It also says the precise site remains controversial and does not establish that the TGN specifically is a major site of production. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites Toh et al., 2017 sourcePartly supported

  The Discussion reports that blocking APP export from the TGN increases Aβ production and suggests that Aβ can be generated there. It calls the TGN a potentially important site, but does not establish that it is a major site of Aβ production along the secretory pathway under normal conditions. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

GGA1 is not required for post-Golgi export of BACE1.

“In contrast, GGA1, which is also found at the Golgi, is not required for the post-Golgi export of BACE1.”

• cites He et al., 2002Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites He et al., 2005Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Toh et al., 2018 sourceCould not verify

  The Results section on GGA1 depletion and the Discussion address BACE1 transport from early endosomes to recycling endosomes, not its export from the Golgi. A search for post-Golgi export and Golgi-to-plasma-membrane trafficking found no passage in our copy establishing whether GGA1 is required for that step. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

Depletion of Arf1 or Arf4 does not affect recruitment of AP-4 to the TGN.

“We have previously shown that depletion of Arf1 or Arf4 does not affect recruitment of AP-4 to the TGN (Toh et al., 2017).”

• cites Toh et al., 2017 sourceSupported

There were low levels of BACE1-SBP-GFP (10.2 ± 2.3%) in CD63-positive late endosome/lysosome at both 45 and 60 min.

“There were low levels of BACE1-SBP-GFP (10.2 ± 2.3%) in CD63-positive late endosome/lysosome at both the 45- and 60-min time points”

AP-1-depleted cells showed a 35% reduction in the slope of Golgi exit compared with control and AP-4 siRNA-treated cells.

“AP-1–depleted cells showed a 35% reduction in slope compared with control and AP-4 siRNA-treated cells (Figure 7E).”

Colocalization of BACE1-GFP and golgin-97 increased from 8.1 ± 2.1% in control cells to 21.1 ± 2.4% in Arf1-depleted cells and 25.7 ± 2.9% in Arf4-depleted cells.

“Quantitation revealed that there was an increase in the colocalization of BACE1-GFP and golgin-97 from 8.1 ± 2.1% in control cells to 21.1 ± 2.4% and 25.7 ± 2.9% in Arf1- and Arf4-depleted cells, respectively (Figure 8D).”

Secreted Aβ increased 1.7-fold from AP-1γ-depleted cells compared with control.

“There was a 1.7-fold increase in secreted Aβ from AP-1γ–depleted cells as compared with control cells (Figure 10A).”

Cell surface expression of BACE1 was unaltered in AP-4ε- or Arl5b-depleted cells.

“cell surface expression of BACE1 was unaltered in either AP-4ε− or Arl5b-depleted cells (Figure 9, B and E).”

In Arf1+Arf4-depleted cells, BACE1-GFP accumulated in the TGN to a lesser extent than after single depletion of Arf4.

“In Arf1+Arf4–depleted cells, BACE1-GFP accumulated in the TGN, as expected, but to a lesser extent than after single depletion of Arf4 (Figure 8, B and C).”

Depletion of AP-1γ results in a decrease in the total cell pool of BACE1-GFP that overlapped with Rab11.

“Conversely, the depletion of AP-1γ resulted in a decrease in the total cell pool of BACE1-GFP that overlapped with Rab11 (Figure 5)”

BACE1-SBP-GFP in the TGN decreased to 15.1 ± 2.5% at 45 min and 11.7 ± 2.2% at 60 min.

“the level of BACE1-SBP-GFP in the TGN decreased to 15.1 ± 2.5% at 45 min and 11.7 ± 2.2% at 60 min”

Depletion of either AP-4 or Arl5b did not alter the distribution of BACE1-GFP at the Golgi.

“As expected, the depletion of either AP-4 or Arl5b in HeLa-BACE1-GFP also did not alter the distribution of BACE1-GFP at the Golgi (Figure 4, A–C).”

Depletion of AP-1γ showed no significant change in cell surface TfR compared with control.

“the depletion of AP-1γ showed no significant change in the level of cell surface TfR from that in control cells (Supplemental Figure S6, A and C).”

Newly synthesized BACE1 traffics directly from the TGN to the plasma membrane.

“We have demonstrated that newly synthesized BACE1 traffics from the TGN directly to the cell surface.”

• Also stated in: § Trafficking itinerary of newly synthesized BACE1 from the TGN · hedged

  “Taken together, TIRF and fast-acquisition imaging data indicate that newly synthesized BACE1 traffics directly from the TGN to the PM.”

BACE1 mean fluorescence intensity decreased by 36.3% and 39.3% in Arf1 siRNA and Arf4 siRNA treated cells, respectively, compared with control.

“Quantitative analysis showed a very similar decrease in BACE1 mean fluorescence intensity by 36.3% and 39.3% in cells treated with Arf1 siRNA or Arf4 siRNA, respectively, compared with control conditions (Figure 9F).”

BACE1 is transported from the trans-Golgi network to the plasma membrane in an AP-1- and Arf1/4-dependent manner.

“we show that BACE1 is transported from the trans-Golgi network to the plasma membrane in an AP-1- and Arf1/4-dependent manner.”

Depletion of AP-1γ, Arf1, or Arf4 did not alter the localization of APP at the TGN compared with control siRNA.

“None of these knockdowns altered the localization of APP at the TGN from that in cells treated with control siRNA, as shown by quantitative analyses (Supplemental Figure S5).”

In primary mouse cortical neurons, AP-1 depletion increases colocalization of endogenous BACE1 with the TGN marker p230/golgin-245 from 10.4 ± 3.1% to 25.2 ± 5.6%.

“Quantitative analyses of fluorescence intensities of 3D projections revealed an increase in colocalization of BACE1 with the TGN marker p230/golgin-245, from 10.4 ± 3.1% in nontransduced neurons to 25.2 ± 5.6% in AP-1–depleted neurons (Figure 6B).”

AP-1γ-depleted HeLa-BACE1-GFP cells showed a reduction of cell surface BACE1 compared with control.

“AP-1γ–depleted HeLa-BACE1-GFP cells showed a reduction of cell surface BACE1 compared with control conditions (Figure 9A).”

Depletion of Arf4 resulted in a greater accumulation of BACE1-GFP at the TGN than depletion of Arf1.

“Notably, the depletion of Arf4 resulted in a greater accumulation of BACE1-GFP at the TGN than the depletion of Arf1 (Figure 8D).”

Quantitative analyses revealed a 3.0-fold increase in colocalization of BACE1-GFP with the TGN marker golgin-97 in AP-1γ-depleted cells compared with control siRNA-treated cells.

“Quantitative analyses revealed a 3.0-fold increase in colocalization of BACE1-GFP with the TGN marker golgin-97 in AP-1γ–depleted cells compared with control siRNA-treated cells (Figure 4D).”

Depletion of AP-1γ in neurons did not alter the distribution of APP in the Golgi.

“depletion of AP-1γ in neurons did not alter the distribution of APP in the Golgi (Figure 6, C and D).”

BACE1-GFP did not accumulate in the Golgi of cells expressing either the Arf1 or Arf4 rescue construct, confirming that BACE1-GFP accumulation in the TGN was specifically due to depletion of Arf1 and Arf4.

“BACE1-GFP did not accumulate in the Golgi of cells expressing either the Arf1 and Arf4 rescue construct, confirming that the accumulation of BACE1-GFP in the TGN was specifically due to the depletion of Arf1 and Arf4 (Figure 8, B, E, and F).”

BACE1 exits the Golgi in HeLa cells and primary neurons by a pathway distinct from the trafficking pathway for APP.

“Here we demonstrate that BACE1 exits the Golgi in HeLa cells and primary neurons by a pathway distinct from the trafficking pathway for APP.”

BACE1 mean fluorescence intensity decreased by 19.8% in AP-1γ-depleted cells compared with control siRNA-treated cells.

“There was a decrease in BACE1 mean fluorescence intensity of 19.8% in AP-1γ–depleted cells compared with control siRNA-treated cells (Figure 9D)”

AP-4 depletion does not significantly change the colocalization of BACE1 with the TGN marker in primary neurons.

“following transduction of primary neurons with AP-4ε shRNA recombinant lentivirus (Supplemental Figure S4B), there was no significant change in the colocalization of BACE1 with the TGN marker in AP-4–depleted neurons (Figure 6B).”

The Golgi exit of BACE1-SBP-GFP was delayed in AP-1-depleted cells compared with control cells and cells treated with AP-4ε siRNA.

“From these findings, we conclude that the Golgi exit of BACE1-SBP-GFP was delayed in AP-1–depleted cells compared with control cells and cells treated with AP-4ε siRNA”

After 10 min of biotin addition, 11.5 ± 3.5% of BACE1-SBP-GFP colocalized with the TGN marker GCC88.

“After 10 min incubation with biotin, BACE1-SBP-GFP exhibited a juxtanuclear staining pattern with very little overlap with the ER marker KDEL (Supplemental Figure S1A) or endosomal markers (Figure 1C; Supplemental Figure S1C) and with 11.5 ± 3.5% of BACE1-SBP-GFP colocalized with the TGN marker GCC88 (Figure 1C; Supplemental Figure S1D)”

A previously uncharacterized acidic cluster-sorting signal, DDFADDISLL, exists in the cytoplasmic tail of BACE1.

“we identified a previously uncharacterized acidic cluster–sorting signal in the cytoplasmic tail of BACE1.”

• Also stated in: § AP-1 is required for the post-Golgi sorting of BACE1 in HeLa cells · plain

  “In the cytoplasmic tail of BACE1, we came across a previously undetected acidic cluster sequence, DDFADDISLL, a motif presenting an extension of the previously defined DDISLL and DISLL motifs that play a role in endocytosis and endocytic sorting (He et al., 2002, 2005; Toh et al., 2018).”

BACE1 is endocytosed to early and recycling endosomes.

“Subsequently, BACE1 is endocytosed to early and recycling endosomes.”

AP-1 and the Arf small G proteins Arf1 and Arf4 are required for efficient Golgi export of BACE1 in both HeLa cells and primary neurons.

“We have demonstrated that AP-1 and the Arf small G proteins Arf1 and Arf4 are required for efficient Golgi export of BACE1 in both HeLa cells and primary neurons.”

Depletion of either Arf1 or Arf4 accumulates BACE1-GFP in the juxtanuclear region that colocalizes extensively with the Golgi marker golgin-97 compared with control conditions.

“The depletion of either Arf1 or Arf4 accumulated BACE1-GFP in the juxtanuclear region that colocalizes extensively with the Golgi marker golgin-97, compared with control conditions (Figure 8B).”

BACE1-GFP colocalization with golgin-97 increased from 8% in control cells to 17.1 ± 4.8% in Arf1+Arf4-depleted cells.

“Quantitation revealed an increase in BACE1-GFP colocalization with golgin-97 from 8% in control cells to 17.1 ± 4.8% in Arf1+Arf4–depleted cells (Figure 8D).”

Depletion of either Arf1 or Arf4 in HeLa-BACE1-GFP resulted in decreased cell surface expression of BACE1.

“The depletion of either Arf1 or Arf4 in HeLa-BACE1-GFP resulted in a decrease in cell surface expression of BACE1, as shown by a leftward shift in the fluorescence intensity peaks of each knockdown condition from control conditions (Figure 9C).”

The level of BACE1-GFP at the TGN was not altered in GGA1-depleted cells.

“On the other hand, the level of BACE1-GFP at the TGN was not altered in GGA1-depleted cells (Figure 4, C and D).”

AP-1γ was depleted by 43-46% in neurons transduced with AP-1γ-shRNA1 and AP-1γ-shRNA2 lentivirus compared with nontransduced neurons.

“AP-1γ was depleted by 43–46% in neurons transduced with AP-1γ-shRNA1 and AP-1γ-shRNA2 lentivirus as compared with nontransduced neurons (Figure 11, A and B).”

In AP-1-silenced neurons, there was a 3.0-fold increase for Aβ40 and a 3.7-fold increase for Aβ42.

“In AP-1–silenced neurons, there was an increase in levels of both Aβ species: a 3.0-fold increase for Aβ40 and a 3.7-fold increase for Aβ42.”

In absence of DAPT, β-CTF/C99 and α-CTF/C83 were not detected in any neuronal extracts.

“In the absence of DAPT, β-CTF/C99 and α-CTF/C83 were not detected in any of the neuronal extracts (Figure 11A)”

Newly synthesized BACE1 traffics from the TGN to the PM.

“newly synthesized BACE1 traffics from the TGN to the PM”

• Also stated in: § DISCUSSION · plain

  “BACE1-SBP-GFP was transported to the Golgi within 10–15 min following release from the ER by addition of biotin, and then trafficked from the TGN to the PM.”

Depletion of either Arf1 or Arf4 resulted in accumulation of BACE1-GFP in the TGN.

“The depletion of either Arf1 or Arf4 resulted in the accumulation of BACE1-GFP in the TGN.”

Depletion of AP-1, Arf1, or Arf4 resulted in reduction of BACE1 at cell surface.

“Depletion of AP-1, Arf1, or Arf4 resulted in a reduction of BACE1 at the cell surface.”

No significant changes in α-CTF/C83 levels in AP-1γ-shRNA1- and AP-1γ-shRNA2-transduced neurons compared with nontransduced neurons.

“there were no significant changes in the level of α-CTF/C83 (** band) in AP-1γ-shRNA1– and AP-1γ-shRNA2–transduced neurons from that in nontransduced neurons (Figure 11, A and D).”

In presence of DAPT, intracellular β-CTF/C99 increased 2.7-fold, 3.9-fold, and 4.1-fold in Arf1 siRNA, Arf4 siRNA, and Arf1+4 siRNAs treated cells, respectively, compared with control.

“In the presence of DAPT, there was also a 2.7-fold, 3.9-fold, and 4.1-fold increase in the level of intracellular β-CTF/C99 detected in cells treated with either Arf1 siRNA, Arf4 siRNA, or Arf1+4 siRNAs, respectively, from that in control siRNA-treated cells (Figure 10, D and E).”

In presence of C3 and DAPT, the 11-kDa band was absent, confirming its identity as β-CTF/C99.

“In the presence of C3 and DAPT, the 11-kDa band (indicated by *) was absent (Figure 11A), confirming their identity as β-CTF/C99.”

• AssumptionRests on an assumption: The BACE1 inhibitor C3 specifically inhibits BACE1 without affecting other proteases.

Only low levels of β-CTF/C99 were detectable in AP-1-/Arf1-/Arf4-depleted cells in absence of γ-secretase inhibitor.

“Only low levels of β-CTF/C99 were detectable in AP-1-/Arf1-/Arf4-depleted cells in the absence of the γ-secretase inhibitor”

Aβ42 is generated via TGN-mediated processing of APP.

“the finding that Aβ42 is generated via TGN-mediated processing of APP is an important observation”

BACE1-SBP-GFP-containing vesicles were transported from a juxtanuclear compartment directly to the distal edge of the cell.

“quantitative analysis of confocal live imaging, TIRF analysis, and fast image acquisition, which showed that BACE1-SBP-GFP–containing vesicles were transported from a juxtanuclear compartment directly to the distal edge of the cell.”

Upon depletion of AP-1, Arf1, or Arf4, there was increased processing of APP, with 2.8- to 4.1-fold increase of intracellular β-CTF/C99 and 1.7- to 4.5-fold increase in secreted Aβ.

“Upon the depletion of either AP-1, Arf1, or Arf4, there was increased processing of APP, as demonstrated by a 2.8- to 4.1-fold increase of intracellular β-CTF/C99 and a 1.7- to 4.5-fold increase in secreted Aβ secretion compared with control conditions.”

In presence of DAPT, intracellular β-CTF/C99 increased 1.88- and 1.94-fold in neurons transduced with AP-1γ-shRNA1 and AP-1γ-shRNA2, respectively, compared with nontransduced neurons.

“in the presence of DAPT, there were 1.88- and 1.94-fold increases in the level of intracellular β-CTF/C99 in neurons transduced with AP-1γ-shRNA1 and AP-1γ- shRNA2, respectively, compared with nontransduced neurons (Figure 11, A and C).”

Greater increase in β-CTF/C99 was detected under Arf4-depleted conditions than under Arf1-depleted conditions.

“Notably, a greater increase in the level of β-CTF/C99 was detected under Arf4-depleted conditions than under Arf1-depleted conditions (Figure 10, D and E)”

BACE1 was retained in the Golgi/TGN following AP-1 knockdown.

“BACE1 was retained in the Golgi/TGN following AP-1 knockdown, as revealed by localization studies of fixed cells with defined markers and by the analysis of the kinetics of BACE1 transport through the Golgi in live cells using RUSH.”

Golgi exit of BACE1 and APP in primary neurons is tightly regulated, resulting in their segregation along different transport routes, which limits APP processing.

“These findings reveal that Golgi exit of BACE1 and APP in primary neurons is tightly regulated, resulting in their segregation along different transport routes, which limits APP processing.”

BACE1 is the primary β-secretase that initiates β-site cleavage of APP to generate the membrane-bound C99 fragment.

“BACE1 is the primary β-secretase for initiating the β-site cleavage of amyloid precursor protein (APP) to generate the membrane-bound C99 fragment (Hussain et al., 1999; Sinha et al., 1999; Yan et al., 1999; Bennett et al., 2000).”

• cites Hussain et al., 1999 sourceNot checked — no full text of the source
• cites Sinha et al., 1999 sourceNot checked — no full text of the source
• cites Yan et al., 1999 sourceNot checked — no full text of the source

TIRF and fast-acquisition imaging data indicate that newly synthesized BACE1 traffics directly from the TGN to the plasma membrane.

“Taken together, TIRF and fast-acquisition imaging data indicate that newly synthesized BACE1 traffics directly from the TGN to the PM.”

Immunoblotting showed that AP-1γ and GGA1 levels were depleted by >90% and >80%, respectively, in HeLa-BACE1-GFP treated with AP-1γ or GGA1 siRNA.

“Immunoblotting showed that the levels of AP-1γ and GGA1 were depleted by >90% and >80%, respectively, in HeLa-BACE1-GFP treated with either AP-1γ or GGA1 small interfering RNA (siRNA) (Figure 4A).”

The AP-1/Arf1/Arf4 sorting machinery is not required for APP export from the TGN.

“These data indicate that the AP-1/Arf1/Arf4 sorting machinery is not required for APP export from the TGN.”

APP processing requires convergence of BACE1 and APP to the same compartment.

“as both BACE1 and APP are membrane proteins, APP processing requires the convergence of these proteins to the same compartment (Kinoshita et al., 2003).”

• cites Kinoshita et al., 2003 sourceNot checked — no full text of the source

Cell surface levels of BACE1 are dependent on AP-1 mediated trafficking of BACE1 from the Golgi.

“Therefore, the cell surface levels of BACE1 are dependent on AP-1 mediated trafficking of BACE1 from the Golgi.”

Cell surface expression of BACE1 is dependent on the Golgi AP-1 export pathway.

“These results show that cell surface expression of BACE1 is dependent on the Golgi AP-1 export pathway.”

Segregation of BACE1 and APP into distinct pathways at the TGN is relevant to the regulation of APP processing and Aβ production.

“Our findings also demonstrate that the segregation of BACE1 and APP into distinct pathways at the TGN is relevant to the regulation of APP processing and Aβ production.”

Arf1 and Arf4 are required for trafficking of BACE1 to the cell surface.

“The decrease in cell surface expression of BACE1 demonstrates that Arf1 and Arf4 are also required for the trafficking of BACE1 to the cell surface.”

Redirecting the BACE1 plasma membrane recycling pathway via the TGN using a chimeric BACE1/TGN38 construct leads to increased Aβ production.

“redirecting the BACE1 plasma membrane recycling pathway via the trans-Golgi network (TGN), using a chimeric BACE1/TGN38 construct (TGN38 is known to traffic from the endosomes to the TGN), leads to increased Aβ production (Chia et al., 2013).”

• cites Chia et al., 2013 sourceNot checked — no full text of the source

Increase in Aβ production indicates enhanced processing of APP by endogenous BACE1 at the TGN following depletion of AP-1, Arf1, and Arf4 sorting machinery.

“The increase in Aβ production indicates enhanced processing of APP by endogenous BACE1 at the TGN following the depletion of AP-1, Arf1, and Arf4 sorting machinery.”

BACE1-SBP-GFP detected in early endosomes at 30 min likely represents arrival of internalized BACE1 from the plasma membrane.

“Rather, the BACE1-SBP-GFP detected in the early endosomes at 30 min likely represents the arrival of internalized BACE1 from the PM.”

Enhanced APP cleavage was detected at TGN upon redirection of BACE1 recycling to this compartment using BACE1/TGN38 chimera.

“our earlier study where enhanced APP cleavage was detected at the TGN upon the redirection of BACE1 recycling to this compartment using a BACE1/TGN38 chimera (Chia et al., 2013).”

• cites Chia et al., 2013 sourceNot checked — no full text of the source

Post-Golgi trafficking of APP is independent of the AP-1-/Arf1-/Arf4-regulated pathway.

“the post-Golgi trafficking of APP is independent of the AP-1-/Arf1-/Arf4-regulated pathway”

Increase in convergence between BACE1 and APP in TGN leads to increased processing of APP by BACE1 in this compartment.

“The increase in convergence between BACE1 and APP in the TGN then leads to increased processing of APP by BACE1 in this compartment.”

AP-1 pathway is critical for regulating processing events under physiological conditions.

“These findings demonstrate that the AP-1 pathway is critical for regulating processing events under physiological conditions.”

Requirement for distinct transport machinery for post-Golgi transport of APP and BACE1 provides a mechanism to sort and segregate APP and BACE1 into separate pathways.

“The requirement for distinct transport machinery for post-Golgi transport of APP and BACE1 provides a mechanism to sort and segregate APP and BACE1 into separate pathways.”

Cargo that interacts selectively with AP-1 at TGN may be transported from Golgi directly to PM.

“cargo that interacts selectively with AP-1 at the TGN may be transported from the Golgi directly to the PM.”

Liberation of Aβ peptide from β-CTF/C99 by γ-secretase is likely to occur in the TGN.

“the liberation of Aβ peptide from β-CTF/C99 by γ-secretase is also likely to occur in the TGN.”

Arrival of BACE1 at early and recycling endosomes at 30 min most likely represents internalized BACE1 from the PM.

“Arrival of BACE1 at the early and recycling endosomes at the 30-min time point most likely represents internalized BACE1 from the PM (Chia et al., 2013).”

• cites Chia et al., 2013 sourceNot checked — no full text of the source

Combination of AP-1 and GGA is likely to regulate sorting of cargoes to endosomes from TGN.

“These studies suggest that the combination of AP-1 and GGA is likely to regulate the sorting of cargoes to the endosomes from the TGN.”

Golgi-associated APP processing events in primary neurons result in generation of cytotoxic Aβ42 species.

“Therefore, the Golgi-associated APP processing events in primary neurons result in the generation of the cytotoxic Aβ42 species.”

Trafficking of BACE1 to PM via AP-1 pathway may serve as regulatory mechanism to segregate BACE1 from APP at TGN, limiting amyloidogenic processing of APP in this compartment.

“The trafficking of BACE1 to the PM via the AP-1 pathway may serve as a regulatory mechanism to segregate BACE1 from APP at the TGN, thus limiting the amyloidogenic processing of APP in this compartment.”

AP-1, Arf1, and Arf4 trafficking machinery is relevant to regulation of amyloidogenic processing of APP at the TGN.

“Collectively, these findings highlight the relevance of AP-1, Arf1, and Arf4 trafficking machinery on the regulation of the amyloidogenic processing of APP at the TGN.”

Trafficking of APP from TGN is independent of AP-1/Arf1/Arf4 pathway.

“the trafficking of APP from the TGN is independent of this pathway.”

AP-1 is required for efficient post-Golgi sorting of BACE1 in both HeLa cells and neurons.

“A key finding of this work is that AP-1 is required for the efficient post-Golgi sorting of BACE1 in both HeLa cells and neurons.”

BACE1 is transported directly from the TGN to the PM.

“Our data strongly indicate that BACE1 is transported directly from the TGN to the PM.”

γ-secretase is localized in the Golgi compartment.

“γ-secretase is also localized in the Golgi compartment (Rechards et al., 2003)”

• cites Rechards et al., 2003 sourceNot checked — no full text of the source

Trafficking of APP from TGN is regulated by AP-4/Arl5b transport machinery directly to endosomal pathway.

“the trafficking of APP from the TGN is regulated by the AP-4/Arl5b transport machinery directly to the endosomal pathway.”

Newly synthesized APP is transported from the TGN directly to early endosomes by an AP-4-dependent pathway.

“We previously showed that newly synthesized APP is transported from the TGN directly to the early endosomes by an AP-4–dependent pathway.”

γ-secretase processes β-CTF/C99 rapidly.

“indicating that γ-secretase processes β-CTF/C99 rapidly.”

AP-1, Arf1, and Arf4 are essential for TGN-to-PM trafficking of BACE1.

“the AP complex, AP-1, and Arf1 and Arf4 small G proteins are essential for TGN-to-PM trafficking of BACE1”

Post-Golgi sorting machinery and trafficking pathways that regulate APP and BACE1 differ.

“These findings further support the conclusion that the post-Golgi sorting machinery and trafficking pathways that regulate APP and BACE1 differ.”

Post-Golgi transport pathways of BACE1 are common between HeLa cells and primary neurons.

“Confirmation of the findings in HeLa cells by the behavior of endogenous BACE1 in primary neurons demonstrates that the post-Golgi transport pathways of BACE1 are common between the cell types.”

Depletion of AP1γ, Arf1, Arf4, or Arf1+4 enhances BACE1 cleavage of APP at the TGN.

“The increase in β-CTF/C99 production directly demonstrates that the depletion of AP1γ, Arf1, Arf4, or Arf1+4 enhances BACE1 cleavage of APP at the TGN.”

Golgi export of endogenous BACE1 by AP-1 pathway is essential for regulation of amyloidogenic processing.

“Using stably transfected HeLa-APP695wt cells, we showed that Golgi export of endogenous BACE1 by the AP-1 pathway is essential for the regulation of amyloidogenic processing.”

A functional AP-1/Arf1/Arf4 pathway is required for export of BACE1 from Golgi directly to PM.

“In conclusion, a functional AP-1/Arf1/Arf4 pathway is required for the export of BACE1 from the Golgi directly to the PM.”

Arf4 is likely the major small G protein regulating post-Golgi trafficking of BACE1.

“These results again indicate that Arf4 is likely to be the major small G protein regulating the post-Golgi trafficking of BACE1.”

Depletion of either AP-1, Arf1, or Arf4 enhanced amyloidogenic processing of APP.

“the depletion of either AP-1, Arf1, or Arf4 enhanced the amyloidogenic processing of APP”

Reduced levels of BACE1 at cell surface in AP-1 knockdown cells are consistent with retention of BACE1 in the Golgi.

“the reduced levels of BACE1 at the cell surface in AP-1 knockdown cells, as assessed by FACS of nonpermeabilized cells, are also consistent with the retention of BACE1 in the Golgi.”