Journal: Frontiers in Immunology

Article Title: Expression of HSPA14 in patients with acute HIV-1 infection and its effect on HIV-1 replication

doi: 10.3389/fimmu.2023.1123600

Figure Lengend Snippet: List of the antibodies used for immunoblotting and immunoprecipitation assays.

Article Snippet: The overexpression plasmids of HSPA14 (GV657 vector) have been provided by Genechem, China .

Techniques: Western Blot, Immunoprecipitation, Concentration Assay

Journal: Frontiers in Immunology

Article Title: Expression of HSPA14 in patients with acute HIV-1 infection and its effect on HIV-1 replication

doi: 10.3389/fimmu.2023.1123600

Figure Lengend Snippet: List of the primers used for qRT-PCR analysis.

Article Snippet: The overexpression plasmids of HSPA14 (GV657 vector) have been provided by Genechem, China .

Techniques: Purification

Journal: Frontiers in Immunology

Article Title: Expression of HSPA14 in patients with acute HIV-1 infection and its effect on HIV-1 replication

doi: 10.3389/fimmu.2023.1123600

Figure Lengend Snippet: Co-IP to detect the interaction between HspBP1 and HSPA14. Co-immunoprecipitation showing interaction of HspBP1 with HSPA14 at endogenous level in infected CEM cells and CD4+T cells of acute HIV infected patients.

Article Snippet: The overexpression plasmids of HSPA14 (GV657 vector) have been provided by Genechem, China .

Techniques: Co-Immunoprecipitation Assay, Immunoprecipitation, Infection

Journal: Frontiers in Immunology

Article Title: Expression of HSPA14 in patients with acute HIV-1 infection and its effect on HIV-1 replication

doi: 10.3389/fimmu.2023.1123600

Figure Lengend Snippet: HSPA14 is down modulated during HIV-1 infection. (A, D) The expression of viral capsid protein p24 mRNA was measured to monitor the progression of infection. (B) mRNA expression profiling of HSPA14 during HIV infection. Different cells were infected with HIV pseudovirus and were harvested on day 1 (D1), day 3 (D3), day 5 (D5), day 7 (D7) and day 9 (D9), post infection for quantitative real time PCR analysis. (C) Representative western blot and densitometry analysis showing expression profile of HSPA14 in HIV pseudovirus infected cells at different time points. (E) mRNA expression profiling of HSPA14 during HIV infection. Different cells were infected with HIV pseudovirus and were harvested on day 1 (D1), day 3 (D3), day 5 (D5) and day 7 (D7), post infection for quantitative real time PCR analysis. (F) Representative western blot and densitometry analysis showing expression profile of HSPA14 in HIV pseudovirus infected CD4+T cells at different time points. CD4+T cells were isolated from PBMC of three healthy donors and were infected with HIV pseudovirus. The error bars are presented as the mean ± SD values and significance is defined as *P ≤ 0.05, **P ≤ 0.01 and ***P ≤ 0.001. INF: Infected with HIV-1. UN, Uninfected.

Article Snippet: The overexpression plasmids of HSPA14 (GV657 vector) have been provided by Genechem, China .

Techniques: Infection, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Isolation

Journal: Frontiers in Immunology

Article Title: Expression of HSPA14 in patients with acute HIV-1 infection and its effect on HIV-1 replication

doi: 10.3389/fimmu.2023.1123600

Figure Lengend Snippet: HSPA14 can interfere the HIV-1 replication (A) Jurkat cells were transfected with indicated plasmids. 24 hours post-transfection, cells were infected with HIV pseudovirus. 24 hours post-infection, cells were harvested to isolate total RNA and protein, and culture supernatants were collected. Upper left panel: qRT-PCR analysis showing different dose of HSP14 over-expression in Jurkat cells. Upper right panel: Culture supernatants were examined for amount of virus produced using p24 antigen capture ELISA. Middle left panel: qRT-PCR analysis revealing change of p24 mRNA levels. Middle right panel and lower panel: representative western blot and densitometry analysis revealing change of p24 protein levels. (B) CEM cells were transfected with indicated plasmids as Jurkat cells. Upper left panel: qRT-PCR analysis showing different dose of HSP14 over-expression in CEM cells. Upper right panel: Culture supernatants were examined for amount of virus produced using p24 antigen capture ELISA. Middle left panel: qRT-PCR analysis revealing change of p24 mRNA levels. Middle right panel and lower panel: representative western blot and densitometry analysis revealing change of p24 protein levels. (C) Jurkat cells were transfected with indicated RNAi. 24 hours post-transfection, cells were infected with HIV pseudovirus. 24 hours post-infection, cells were harvested to isolate total RNA and protein, and culture supernatants were collected. Upper left panel: qRT-PCR analysis showing different dose of HSP14 knockdown in Jurkat cells. Upper right panel: Culture supernatants were examined for amount of virus produced using p24 antigen capture ELISA. Middle left panel: qRT-PCR analysis revealing change of p24 mRNA levels. Middle right panel and lower panel: representative western blot and densitometry analysis revealing change of p24 protein levels. (D) CEM cells were transfected with indicated RNAi as Jurkat cells. Upper left panel: qRT-PCR analysis showing different dose of HSP14 knockdown in CEM cells. Upper right panel: Culture supernatants were examined for amount of virus produced using p24 antigen capture ELISA. Middle left panel: qRT-PCR analysis revealing change of p24 mRNA levels. Middle right panel and lower panel: representative western blot and densitometry analysis revealing change of p24 protein levels. Control: cell lines were transfected with vector plasmids as control. Blank: cell lines were not transfected with any plasmids as blank control. +: 500ng/μl of HSPA14 overexpression plasmid, ++: 1μg/μl of HSPA14 overexpression plasmid, +++: 2μg/μl of HSPA14 overexpression plasmid. -: 50nM of HSPA14 shRNA plasmi, –: 100nM of HSPA14 shRNA plasmid, —: 200nM of HSPA14 shRNA plasmid.The error bars are presented as the mean ± SD values and significance is defined as *P ≤ 0.05, **P ≤ 0.01 and ***P ≤ 0.001.

Article Snippet: The overexpression plasmids of HSPA14 (GV657 vector) have been provided by Genechem, China .

Techniques: Transfection, Infection, Quantitative RT-PCR, Over Expression, Produced, Enzyme-linked Immunosorbent Assay, Western Blot, Plasmid Preparation, shRNA

Journal: Frontiers in Immunology

Article Title: Expression of HSPA14 in patients with acute HIV-1 infection and its effect on HIV-1 replication

doi: 10.3389/fimmu.2023.1123600

Figure Lengend Snippet: Different expression levels of HSPAs in HVL and LVL patients. CD4+ T cells from PBMC of 70 patients were isolated and harvested for RNA and protein lysate preparation. (A) Analyzing and comparing the different HSPA isoforms (including HSPA2, HSPA5, HSPA6, HSPA7, HSPA8, HSPA9, HSPA13 and HSPA14) transcription levels between the LVL and HVL patients by qRT-PCR. (B) Comparing the different HSPA isoforms protein levels between the LVL and HVL patients by western blot and densitometry analysis. Significance is defined as NS: P>0.05, *P ≤ 0.05, **P ≤ 0.01 and ***P ≤ 0.001.

Article Snippet: The overexpression plasmids of HSPA14 (GV657 vector) have been provided by Genechem, China .

Techniques: Expressing, Isolation, Quantitative RT-PCR, Western Blot