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isoform-specific pp1 antibodies α-pp1α/β/γ  (Santa Cruz Biotechnology)


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    Structured Review

    Santa Cruz Biotechnology isoform-specific pp1 antibodies α-pp1α/β/γ
    Isoform Specific Pp1 Antibodies α Pp1α/β/γ, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/pp1+antibodies+%CE%B1+pp1%CE%B1+%CE%B2/anti+pp1%CE%B1/pmc11576475-300-0-8
    Average 90 stars, based on 1 article reviews
    isoform-specific pp1 antibodies α-pp1α/β/γ - by Bioz Stars, 2026-09
    90/100 stars

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    Sequencing:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Co-Immunoprecipitation Assay:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Gene Expression:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Control:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Incubation:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Western Blot:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Comparison:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Transfection:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Two Tailed Test:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    De-Phosphorylation Assay:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Activity Assay:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Expressing:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Phospho-proteomics:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Binding Assay:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Lysis:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Knockdown:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.

    Protein Binding:

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3
    Article Snippet: The membranes were blocked with Tris-buffered saline with 0.1% Tween (TBS-T) containing 5% (wt/vol) non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies.non-fat milk at RT for 60 min. After three washes (5 min), the membranes were incubated overnight at 4°C with the following antibodies in TBS-T with 5% BSA: α-GAPDH (1:1,000, #2118), α-BAG3 (1:1,000, 23842S), and α-PP5 (1:1,000, 2289S), α-CDC37 (1:1,000, 3618S), α-CDC37-pS13 (1:1,000, 13,248), α-phosphoThr (1:500, 42H4, 9386), SQSTM1/p62 (1:1,000, 5114T), and α-14-3-3γ (1:500, 5522S) obtained from Cell Signaling Technologies. ... Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.. The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.The generic α-phospho-Serine (1:500, AB1603) was obtained from Merck.



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    (A) Linear illustration of human BAG3 protein sequence with reported p-sites annotated. (B) Co-IP of FLAG-BAG3 from HEK293 cells revealed significantly enriched proteins, assessed using the rankprod method (n = 4), with a minimum fivefold increase and a P -value ≤ 0.05. Enriched proteins in the sample are marked in respective colors for subcategories. Selected hits are annotated with their gene name. A false discovery rate of 5% is indicated by a dashed line. (C) Enriched phosphatases and respective phosphatase regulators from the BAG3 co-IP sample were assigned to their respective (super-)families of human phosphatases. The phosphoprotein phosphatase family is colored; other Ser/Thr-specific phosphatases are in shades of gray. Gene names are listed next to the corresponding assigned phosphatase identifications. PPMs, metal-dependent protein phosphatases; PSTPs, protein serine/threonine-specific phosphatases. (A, D) Gene ontology enrichment analysis of significantly enriched genes from the BAG3 co-IP sample (n = 277) in (A). The 10 most abundant biological process GO terms are displayed as bars, ranked based on protein counts. Biological processes related to BAG3’s role in protein homeostasis are presented in blue. (E) Gene ontology enrichment analysis of the co-IP sample compared with overall human gene expression, displayed as a heatmap. Fold enrichment was calculated for the BAG3 enriched, not enriched, and total co-IP sets, with overall GO categories annotated at the respective cluster. Statistical significance was determined using Fisher’s test ( P -value < 0.05) with the Bonferroni correction implemented in the PANTHER database. Non-significant changes are colored gray within the heatmap.
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    Image Search Results


    (A) Linear illustration of human BAG3 protein sequence with reported p-sites annotated. (B) Co-IP of FLAG-BAG3 from HEK293 cells revealed significantly enriched proteins, assessed using the rankprod method (n = 4), with a minimum fivefold increase and a P -value ≤ 0.05. Enriched proteins in the sample are marked in respective colors for subcategories. Selected hits are annotated with their gene name. A false discovery rate of 5% is indicated by a dashed line. (C) Enriched phosphatases and respective phosphatase regulators from the BAG3 co-IP sample were assigned to their respective (super-)families of human phosphatases. The phosphoprotein phosphatase family is colored; other Ser/Thr-specific phosphatases are in shades of gray. Gene names are listed next to the corresponding assigned phosphatase identifications. PPMs, metal-dependent protein phosphatases; PSTPs, protein serine/threonine-specific phosphatases. (A, D) Gene ontology enrichment analysis of significantly enriched genes from the BAG3 co-IP sample (n = 277) in (A). The 10 most abundant biological process GO terms are displayed as bars, ranked based on protein counts. Biological processes related to BAG3’s role in protein homeostasis are presented in blue. (E) Gene ontology enrichment analysis of the co-IP sample compared with overall human gene expression, displayed as a heatmap. Fold enrichment was calculated for the BAG3 enriched, not enriched, and total co-IP sets, with overall GO categories annotated at the respective cluster. Statistical significance was determined using Fisher’s test ( P -value < 0.05) with the Bonferroni correction implemented in the PANTHER database. Non-significant changes are colored gray within the heatmap.

    Journal: Life Science Alliance

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3

    doi: 10.26508/lsa.202402734

    Figure Lengend Snippet: (A) Linear illustration of human BAG3 protein sequence with reported p-sites annotated. (B) Co-IP of FLAG-BAG3 from HEK293 cells revealed significantly enriched proteins, assessed using the rankprod method (n = 4), with a minimum fivefold increase and a P -value ≤ 0.05. Enriched proteins in the sample are marked in respective colors for subcategories. Selected hits are annotated with their gene name. A false discovery rate of 5% is indicated by a dashed line. (C) Enriched phosphatases and respective phosphatase regulators from the BAG3 co-IP sample were assigned to their respective (super-)families of human phosphatases. The phosphoprotein phosphatase family is colored; other Ser/Thr-specific phosphatases are in shades of gray. Gene names are listed next to the corresponding assigned phosphatase identifications. PPMs, metal-dependent protein phosphatases; PSTPs, protein serine/threonine-specific phosphatases. (A, D) Gene ontology enrichment analysis of significantly enriched genes from the BAG3 co-IP sample (n = 277) in (A). The 10 most abundant biological process GO terms are displayed as bars, ranked based on protein counts. Biological processes related to BAG3’s role in protein homeostasis are presented in blue. (E) Gene ontology enrichment analysis of the co-IP sample compared with overall human gene expression, displayed as a heatmap. Fold enrichment was calculated for the BAG3 enriched, not enriched, and total co-IP sets, with overall GO categories annotated at the respective cluster. Statistical significance was determined using Fisher’s test ( P -value < 0.05) with the Bonferroni correction implemented in the PANTHER database. Non-significant changes are colored gray within the heatmap.

    Article Snippet: Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.

    Techniques: Sequencing, Co-Immunoprecipitation Assay, Gene Expression

    (A) A7r5 smooth muscle cells were treated with increasing concentrations of phosphoprotein phosphatase inhibitors CalA or OA as indicated, or DMSO as control. After 20 min of incubation, cells were lysed, and modulator effects on BAG3-pS136 were determined with immunoblots. The quantification is presented as a bar plot, with the mean depicted with error bars that represent the SEM based on five independent experiments. Statistical significance between inhibitors was determined with t test and P -values are displayed as stars (* P = 0.020, *** P < 0.001). (B, C) Michaelis–Menten kinetic parameters of PP1c were determined against the indicated mono- or bisphosphorylated peptides. Error bars represent the SEM of three independent replicates with technical duplicates. k cat / K m was calculated by comparison with a phosphate standard curve. (D) Overexpressed FLAG-BAG3 from transiently transfected HEK293 cells was single-step affinity immobilized with anti-FLAG beads and treated with PP1c for 30 min. Immunoblot quantification depicts the mean of four independent experiments, the error bar represents the SD, with P -values obtained from a t tests (two-tailed, paired, **** P < 0.001). (E) Monitoring of BAG3-pS136 dephosphorylation in A7r5 lysate upon incubation with PP1c, PP5, or without phosphatase as a control treatment for the indicated incubation time. Endogenous phosphatases were inhibited by the addition of 20 nM CalA. Quantification depicts the results of four independent experiments with P -values obtained from two-way ANOVA with Sidak correction (** P = 0.006 [PP1c], ** P = 0.005 [PP5], **** P < 0.001). Mean is shown and error bars represent the SEM. Source data are available for this figure.

    Journal: Life Science Alliance

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3

    doi: 10.26508/lsa.202402734

    Figure Lengend Snippet: (A) A7r5 smooth muscle cells were treated with increasing concentrations of phosphoprotein phosphatase inhibitors CalA or OA as indicated, or DMSO as control. After 20 min of incubation, cells were lysed, and modulator effects on BAG3-pS136 were determined with immunoblots. The quantification is presented as a bar plot, with the mean depicted with error bars that represent the SEM based on five independent experiments. Statistical significance between inhibitors was determined with t test and P -values are displayed as stars (* P = 0.020, *** P < 0.001). (B, C) Michaelis–Menten kinetic parameters of PP1c were determined against the indicated mono- or bisphosphorylated peptides. Error bars represent the SEM of three independent replicates with technical duplicates. k cat / K m was calculated by comparison with a phosphate standard curve. (D) Overexpressed FLAG-BAG3 from transiently transfected HEK293 cells was single-step affinity immobilized with anti-FLAG beads and treated with PP1c for 30 min. Immunoblot quantification depicts the mean of four independent experiments, the error bar represents the SD, with P -values obtained from a t tests (two-tailed, paired, **** P < 0.001). (E) Monitoring of BAG3-pS136 dephosphorylation in A7r5 lysate upon incubation with PP1c, PP5, or without phosphatase as a control treatment for the indicated incubation time. Endogenous phosphatases were inhibited by the addition of 20 nM CalA. Quantification depicts the results of four independent experiments with P -values obtained from two-way ANOVA with Sidak correction (** P = 0.006 [PP1c], ** P = 0.005 [PP5], **** P < 0.001). Mean is shown and error bars represent the SEM. Source data are available for this figure.

    Article Snippet: Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.

    Techniques: Control, Incubation, Western Blot, Comparison, Transfection, Two Tailed Test, De-Phosphorylation Assay

    (A) Illustrative overview of the small molecule and peptide modulator screen to tune PP1 activity in cells. HEK293 cells transiently expressing FLAG-BAG3 were treated as illustrated, lysed, and captured by a single-step affinity enrichment using anti-FLAG beads. Samples were analyzed by immunoblots. Quantification depicts results of three independent experiments with P -values obtained from t tests (two-tailed). Mean is shown with replicates as scatter plot and error bars represent the SD (* P = 0.015, *** P < 0.001). (B) A7r5 smooth muscle cells were incubated with increasing concentrations of Tautomycetin as indicated. Immunoblots of lysate were used to determine the sensitivity of the titrated inhibitor towards BAG3-pS136. Mean is shown as a barplot with replicates as scatter plots, and error bars represent the SD of four independent experiments. Statistical significance between concentrations is determined with t test with Welch’s correction (* P < 0.05). (C) Analysis of phosphorylation-dependent binding of 14-3-3γ to FLAG-BAG3 in A7r5 muscle cells after 20-min modulation of endogenous PP1 before lysis and FLAG-IP enrichment. Results are presented as a barplot, P -values obtained from a t test (*** P = 0.0002, **** P < 0.0001). (D) Immunoblots depicting siRNA-mediated knockdown of all PP1 isoforms combined (PP1α/β/γ) or individual PP1 isoforms (PP1α, PP1β, PP1γ) separately. The displayed blots represent the analyzed protein levels of 24 or 48 h after transfection. (E) Quantification of immunoblotted lysates show level changes of PP1α/β/γ and BAG3 (n = 3 or 4). Quantification depicts results of three or four independent experiments with P -values obtained from t tests (two-tailed, paired). Error bars represent the SD. Source data are available for this figure.

    Journal: Life Science Alliance

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3

    doi: 10.26508/lsa.202402734

    Figure Lengend Snippet: (A) Illustrative overview of the small molecule and peptide modulator screen to tune PP1 activity in cells. HEK293 cells transiently expressing FLAG-BAG3 were treated as illustrated, lysed, and captured by a single-step affinity enrichment using anti-FLAG beads. Samples were analyzed by immunoblots. Quantification depicts results of three independent experiments with P -values obtained from t tests (two-tailed). Mean is shown with replicates as scatter plot and error bars represent the SD (* P = 0.015, *** P < 0.001). (B) A7r5 smooth muscle cells were incubated with increasing concentrations of Tautomycetin as indicated. Immunoblots of lysate were used to determine the sensitivity of the titrated inhibitor towards BAG3-pS136. Mean is shown as a barplot with replicates as scatter plots, and error bars represent the SD of four independent experiments. Statistical significance between concentrations is determined with t test with Welch’s correction (* P < 0.05). (C) Analysis of phosphorylation-dependent binding of 14-3-3γ to FLAG-BAG3 in A7r5 muscle cells after 20-min modulation of endogenous PP1 before lysis and FLAG-IP enrichment. Results are presented as a barplot, P -values obtained from a t test (*** P = 0.0002, **** P < 0.0001). (D) Immunoblots depicting siRNA-mediated knockdown of all PP1 isoforms combined (PP1α/β/γ) or individual PP1 isoforms (PP1α, PP1β, PP1γ) separately. The displayed blots represent the analyzed protein levels of 24 or 48 h after transfection. (E) Quantification of immunoblotted lysates show level changes of PP1α/β/γ and BAG3 (n = 3 or 4). Quantification depicts results of three or four independent experiments with P -values obtained from t tests (two-tailed, paired). Error bars represent the SD. Source data are available for this figure.

    Article Snippet: Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.

    Techniques: Activity Assay, Expressing, Western Blot, Two Tailed Test, Incubation, Phospho-proteomics, Binding Assay, Lysis, Knockdown, Transfection

    (A) Schematic illustration of the protein level assessment after treatment of HEK293 cells with pre-designed siRNA. HEK293 cells were transfected with siRNA of PP1 isoforms (PPP1CA/B/C) for 24 and 48 h. (B) Quantification of immunoblotted lysates show phosphorylation level changes of BAG3-pS136 relative to the overall BAG3 levels (n = 3 or 4). Quantification depicts results of three or four independent experiments with P -values obtained from t tests (two-tailed, paired). Error bars represent the SD. Source data are available for this figure.

    Journal: Life Science Alliance

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3

    doi: 10.26508/lsa.202402734

    Figure Lengend Snippet: (A) Schematic illustration of the protein level assessment after treatment of HEK293 cells with pre-designed siRNA. HEK293 cells were transfected with siRNA of PP1 isoforms (PPP1CA/B/C) for 24 and 48 h. (B) Quantification of immunoblotted lysates show phosphorylation level changes of BAG3-pS136 relative to the overall BAG3 levels (n = 3 or 4). Quantification depicts results of three or four independent experiments with P -values obtained from t tests (two-tailed, paired). Error bars represent the SD. Source data are available for this figure.

    Article Snippet: Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.

    Techniques: Transfection, Phospho-proteomics, Two Tailed Test

    Dephosphorylation of pS136 on BAG3 by PP1 leads to loss of 14-3-3 protein binding. Dephosphorylation of the p-site cluster pS284-pS291 by PP5 enables HspB8 binding, and PP5 depletion increases BAG3 levels in a CASA-dependent manner.

    Journal: Life Science Alliance

    Article Title: Identification of phosphatases that dephosphorylate the co-chaperone BAG3

    doi: 10.26508/lsa.202402734

    Figure Lengend Snippet: Dephosphorylation of pS136 on BAG3 by PP1 leads to loss of 14-3-3 protein binding. Dephosphorylation of the p-site cluster pS284-pS291 by PP5 enables HspB8 binding, and PP5 depletion increases BAG3 levels in a CASA-dependent manner.

    Article Snippet: Isoform-specific PP1 antibodies α-PP1α/β/γ (1:1,000, sc-271762/sc-365678/sc-515943) were from Santa Cruz.

    Techniques: De-Phosphorylation Assay, Protein Binding, Binding Assay