anti lamp2  (Boster Bio)

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: (A) Immunohistochemical staining of LAMP2A in gastric tumor and normal tissue. (Scale bar: 20 μm). (B) The TCGA and GEO databases show the mRNA expression levels of LAMP2A in gastric tumors and normal tissue. (C)The qPCR results showed the LAMP2A expression at mRNA levels. (D) Western blot analysis of the level of LAMP2A in several cell lines. Note:TCGA( https://portal.gdc.cancer.gov/)GEO(https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE63089 ). Data are presented as mean±SD of three independent experiments. ns, no significance, *P < 0.05. **P < 0.01. ***P < 0.001.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Immunohistochemical staining, Staining, Expressing, Western Blot

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: Four cell lines were exposed to H 2 O 2 (150μM, 300μM), respectively, to establish a severe oxidative stress cell model. WB results showed the protein expression of LAMP2A. Data are presented as mean±SD of three independent experiments. ns, no significance, *P < 0.05. **P < 0.01.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Expressing

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: (A, B) Verification of LAMP2A knockdown efficiency in MKN45 cell. (C, D) Verification of LAMP2A overexpression efficiency in AGS cell. Note: NC, negative control. Data are presented as mean±SD of three independent experiments. ns, no significance, *P < 0.05. **P < 0.01.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Knockdown, Over Expression, Negative Control

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: (A): Proliferation of the control group and LAMP2A-knockdown group. (B): Proliferation of the control group and LAMP2A-knockdown group following 150 μM H 2 O 2 treatment. (C): Proliferation of the control group and LAMP2A-overexpress group. (D): Proliferation of the control group and LAMP2A-overexpress group following 150 μM H2O2 treatment. (E): LAMP2A knockdown cell lines were exposed to H 2 O 2 (0, 150μM and 300μM), and the apoptosis rate was measured by FACS after 24-hour treatment. (F): LAMP2A over-expressing cells were exposed to H 2 O 2 (0, 150μM and 300μM), and the apoptosis rate was measured by FACS after 24-hour treatment. Note: L2A, LAMP2A; NC, negative control. Data are presented as mean±SD of three independent experiments. ns, no significance, *P < 0.05. **P < 0.01.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Control, Knockdown, Expressing, Negative Control

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: (A): Identification of a conserved KFERQ-like motif in DJ-1, a CMA substrate recognition signature. (B): WB analysis of DJ-1 protein expression levels. (C) H₂O₂ dose-dependent upregulation of DJ-1 protein in gastric cancer cells (150 μM, 300 μM; 24 h treatment). (D): Co-IP confirms enhanced DJ-1/LAMP2A interaction in H₂O₂-treated (300 μM, 24 h) MKN45 cancer cells. (E): IF demonstrates oxidative stress-induced colocalization of DJ-1 and LAMP2A in MKN45 cells (300 μM H₂O₂, 24 h). Data are presented as mean±SD of three independent experiments. ns, no significance, *P < 0.05. **P < 0.01.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Expressing, Co-Immunoprecipitation Assay

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: (A) MKN45shL2A and MKN45shNC were stimulated by H 2 O 2 (0,300 μm) for 24 hours, and the protein levels of LAMP2A, DJ-1, apoptosis-related proteins Bcl-2 and BAX were detected by WB. (B) The MKN45shL2A cells were transfected with the LAMP2A plasmid to generate the rescue group (sh L2A + LAMP2A). The protein levels of LAMP2A, DJ-1, Bcl-2, and BAX were analyzed by WB in all cell groups after a 24-hour exposure to 300 μM H₂O₂. Data are presented as mean±SD of three independent experiments. ns, no significance, *P < 0.05. **P < 0.01.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Transfection, Plasmid Preparation

Journal: PLOS One

Article Title: LAMP2A-dependent chaperone-mediated autophagy enhances oxidative stress resistance in gastric cancer cells through selective degradation of accumulated oxidized DJ-1

doi: 10.1371/journal.pone.0331823

Figure Lengend Snippet: ①CMA inhibits tumor cell apoptosis through selective removal of overoxidized DJ-1 protein. ②In pathological conditions, ROS-mediated overoxidation converts DJ-1 into a pro-apoptotic form that triggers gastric cancer cell death, distinct from its physiological oxidative modification under basal oxidative stress. ③Oxidative stress serves as the primary inducer of LAMP2A upregulation in gastric cancer cells, establishing a compensatory mechanism for CMA activation.

Article Snippet: The lentiviral vectors with shRNAs against LAMP2A were purchased from GeneChem Company, Shanghai, China (PIEL248064052).

Techniques: Modification, Activation Assay

Journal: Nature Communications

Article Title: Lysosome-targeting live attenuated influenza vaccines elicit robust and broad immunity in mice

doi: 10.1038/s41467-026-69920-0

Figure Lengend Snippet: a LTM-dependent degradation of viral NS1 protein. Western blot analysis shows reduced NS1 protein levels in cells expressing NS1-N LTM compared with the N LTM mutant , while NS1 mRNA levels remain comparable ( n = 3). b Lysosome dependence of LTM-mediated NS1 degradation. NS1-N LTM protein degradation is blocked by lysosomal inhibitors but not by proteasome or autophagy inhibition. HEK293T cells expressing either NS1-N LTM (left) or NS1-N LTM mutant (right) protein were cultured with or without the proteasome inhibitor MG-132 (10 µM), the autophagy inhibitor 3-methyladenine (3-MA; 10 mM), bafilomycin A1 (Baf A1; 0.4 µM), or chloroquine (CQ; 50 µM) for 6 h. The viral NS1 protein was detected by Western blotting ( n = 3). c Co-immunoprecipitation demonstrating interaction of HSC70 with NS1-N LTM but not with the NS1 LTM mutant ( n = 3). d Dependence of LTM-mediated NS1 degradation on LAMP2A. Conventional HEK293T cells and LAMP2A-KO HEK293T cells were transfected with constructs expressing NS1-N LTM (left) or NS1-N LTM mutant (right) protein and collected 24 h post-transfection. Viral NS1 protein was detected by Western blotting ( n = 3). Immunofluorescence analysis showing colocalization of NS1-N LTM with HSC70 ( e ) and LAMP2A ( f ), but not of the NS1-N LTM mutant . Green, NS1; red, HSC70 or LAMP2A; blue, nuclei; yellow, colocalization sites; scale bar, 5 µm. Representative images of at least three independent experiments are shown. LAMP2A-dependent degradation of NS1-N LTM during viral infection in conventional and LAMP2A-KO HEK293T ( g ) and A549 ( h ) cells. Replication competence of NS1-N LTM and NS1-N LTM mutant viruses in conventional and LAMP2A-KO HEK293T ( i ) and A549 ( j ) cells. Immunofluorescence staining of influenza viral M1 protein at 48 h after infection (MOI = 0.01) showing the replication competence of NS1-N LTM or NS1-N LTM mutant virus in conventional and LAMP2A-KO cells. Green, M1; blue, nuclei; scale bar, 100 µm. Viral titers in culture supernatants were quantified by immunofluorescence focus-forming unit (FFU) assay ( n = 3). Data are means ± s.d; n = 3 biologically independent experiments; unpaired two-tailed t -test for ( i ) and ( j ); *** P < 0.001. Source data are provided as a Source Data file.

Article Snippet: After another 6 h of culture, the cells were washed with PBS, fixed with 4% paraformaldehyde (PFA) for 20 min at room temperature, washed with PBS, permeabilized with PBS containing 0.1% Triton X-100 for 5 min, blocked with PBS containing with 0.1% Triton X-100 and 10% goat serum for 1 h at room temperature, and incubated with anti-Flag antibody (MBL, Cat# PM020; 1:1000 dilution) and anti-HSC70 antibody (Santa Cruz Biotechnology, Cat# sc-7298; 1:300 dilution) or anti-LAMP2A antibody (Santa Cruz Biotechnology, Cat# sc-18822; 1:300 dilution) diluted in PBS containing 0.1% Triton X-100 and 1% goat serum overnight at 4 °C, followed by incubation with Alexa Fluor 488 goat anti-rabbit IgG (Life Technologies, Cat# A11034; 1:1000 dilution) and Alexa Fluor 555 goat anti-mouse IgG (Life Technologies, Cat# A32727; 1:1000 dilution) for 1 h at room temperature.

Techniques: Western Blot, Expressing, Mutagenesis, Inhibition, Cell Culture, Immunoprecipitation, Transfection, Construct, Immunofluorescence, Infection, Staining, Virus, Two Tailed Test

Journal: Nature Communications

Article Title: Lysosome-targeting live attenuated influenza vaccines elicit robust and broad immunity in mice

doi: 10.1038/s41467-026-69920-0

Figure Lengend Snippet: a Schematic illustration depicting the design and generation of LYTAR 2.0 influenza viruses. LYTAR 2.0 viruses are attenuated in conventional cells through LTM-mediated viral protein degradation by the CMA system but can replicate efficiently in LAMP2A-knockout (KO) cells. VP stands for viral protein. b Western blots showing the dependence of LTM-mediated viral protein degradation on the lysosome. HEK293T cells expressing the indicated viral proteins were cultured in the presence or absence of Baf A1 (0.4 µM) for 6 h. The viral proteins were detected by Western blotting ( n = 3). c Western blots showing the LAMP2A dependency of LTM-mediated viral protein degradation. Conventional HEK293T cells and LAMP2A-KO HEK293T cells were transfected with constructs expressing the indicated viral proteins and collected 24 h post-transfection for viral protein detection by Western blotting ( n = 3). d Western blots showing the LAMP2A dependency of LTM-mediated viral protein degradation in HEK293T cells. Conventional HEK293T cells and LAMP2A-KO HEK293T cells were infected with the indicated LYTAR 2.0 viruses and collected 48 h post-infection for detection of viral proteins by Western blotting ( n = 3). e Western blots showing the LAMP2A dependence of LTM-mediated viral protein degradation in A549 cells. Conventional A549 cells and LAMP2A-KO A549 cells were infected with the indicated LYTAR 2.0 viruses and collected 48 h post-infection for detection of viral proteins by Western blotting ( n = 3). Source data are provided as a Source Data file.

Article Snippet: After another 6 h of culture, the cells were washed with PBS, fixed with 4% paraformaldehyde (PFA) for 20 min at room temperature, washed with PBS, permeabilized with PBS containing 0.1% Triton X-100 for 5 min, blocked with PBS containing with 0.1% Triton X-100 and 10% goat serum for 1 h at room temperature, and incubated with anti-Flag antibody (MBL, Cat# PM020; 1:1000 dilution) and anti-HSC70 antibody (Santa Cruz Biotechnology, Cat# sc-7298; 1:300 dilution) or anti-LAMP2A antibody (Santa Cruz Biotechnology, Cat# sc-18822; 1:300 dilution) diluted in PBS containing 0.1% Triton X-100 and 1% goat serum overnight at 4 °C, followed by incubation with Alexa Fluor 488 goat anti-rabbit IgG (Life Technologies, Cat# A11034; 1:1000 dilution) and Alexa Fluor 555 goat anti-mouse IgG (Life Technologies, Cat# A32727; 1:1000 dilution) for 1 h at room temperature.

Techniques: Knock-Out, Western Blot, Expressing, Cell Culture, Transfection, Construct, Infection

Journal: Nature Communications

Article Title: Lysosome-targeting live attenuated influenza vaccines elicit robust and broad immunity in mice

doi: 10.1038/s41467-026-69920-0

Figure Lengend Snippet: a Multi-cycle replication kinetics of the indicated viruses in conventional and LAMP2A-KO MDCK cells. Data are presented as means ± s.d ( n = 3). b triLTM-dependent degradation of viral proteins. Western blot analysis shows reduced levels of triLTM-tagged viral proteins compared with mutated triLTM-tagged controls, while corresponding mRNA levels remain comparable ( n = 3). c Lysosome dependence of triLTM-mediated viral protein degradation. HEK293T cells expressing triLTM-tagged or mutated triLTM-tagged viral proteins were cultured in the presence or absence of Baf A1 (0.4 µM) for 6 h and collected for detection of indicated viral proteins by Western blotting ( n = 3). d Co-immunoprecipitation demonstrating interaction of HSC70 with triLTM-tagged viral proteins but not with mutated triLTM-tagged viral proteins ( n = 3). LAMP2A-dependent degradation of triLTM-tagged viral proteins during viral infection in conventional and LAMP2A-KO HEK293T ( e ) and A549 ( f ) cells ( n = 3). Conventional cells and LAMP2A-KO cells were infected with LYTAR 2.0 dual triLTMs or LYTAR 2.0 dual triLTMs mutant virus and collected at 48 h after infection for detection of indicated proteins by Western blotting ( n = 3). Replication competence of LYTAR 2.0 dual triLTMs and LYTAR 2.0 dual triLTMs mutant viruses in conventional and LAMP2A-KO HEK293T ( g ) and A549 ( h ) cells. Immunofluorescence staining of influenza viral M1 protein at 48 h after infection (MOI = 0.01) showing the replication competence of LYTAR 2.0 dual triLTMs and LYTAR 2.0 dual triLTMs mutant virus in conventional and LAMP2A-KO HEK293T cells. Green, M1; blue, nuclei; scale bar, 100 µm. Viral titers in culture supernatants were quantified by immunofluorescence focus-forming unit (FFU) assay ( n = 3). Data are means ± s.d; n = 3 biologically independent experiments; unpaired two-tailed t -test for ( g ) and ( h ); *** P < 0.001. Source data are provided as a Source Data file.

Article Snippet: After another 6 h of culture, the cells were washed with PBS, fixed with 4% paraformaldehyde (PFA) for 20 min at room temperature, washed with PBS, permeabilized with PBS containing 0.1% Triton X-100 for 5 min, blocked with PBS containing with 0.1% Triton X-100 and 10% goat serum for 1 h at room temperature, and incubated with anti-Flag antibody (MBL, Cat# PM020; 1:1000 dilution) and anti-HSC70 antibody (Santa Cruz Biotechnology, Cat# sc-7298; 1:300 dilution) or anti-LAMP2A antibody (Santa Cruz Biotechnology, Cat# sc-18822; 1:300 dilution) diluted in PBS containing 0.1% Triton X-100 and 1% goat serum overnight at 4 °C, followed by incubation with Alexa Fluor 488 goat anti-rabbit IgG (Life Technologies, Cat# A11034; 1:1000 dilution) and Alexa Fluor 555 goat anti-mouse IgG (Life Technologies, Cat# A32727; 1:1000 dilution) for 1 h at room temperature.

Techniques: Western Blot, Expressing, Cell Culture, Immunoprecipitation, Infection, Mutagenesis, Virus, Immunofluorescence, Staining, Two Tailed Test