(A) Bone marrow-derived macrophage (BMDM) from wild-type (WT), p38γ/δKO, and p38γ/δKIKO mice was exposed to LPS (100 ng/ml) for the indicated times. Cell lysates (30 μg) were immunoblotted with the …
Labelled (.pdf) and raw (.jpeg) western blot images showed in panel A.
(A) Samples of genomic DNA from wild-type (WT), Mapk13−/−, Mapk12171A/171A, and p38γ/δKIKO purified from tail biopsy were used as templates for PCR as described in methods. (B) WT and p38γ/δKIKO …
Labelled (.pdf) and raw (.tif) western blot images showed in panel A.
Labelled (.pdf) and raw (.tif,. jpeg) western blot images showed in panel B.
Labelled (.pdf) and raw (.jpeg) western blot images showed in panel C.
Labelled (.pdf) and raw (.jpeg) western blot images showed in panel D.
Labelled (.pdf) and raw (.jpeg) western blot images showed in panel E.
(A) Wild-type (WT) and p38γ/δKIKO mice were intravenously injected with 1 × 105 CFU of C. albicans. Kidney fungal load was determined 3 days after infection. Each symbol represents an individual …
(A) In vitro bone marrow-derived macrophage (BMDM) development is not affected in p38γ/δKIKO mice. BMDM from wild-type (WT) and p38γ/δKIKO mice was stained with anti-F4/80 antibody, and analysed by …
(A) Bone marrow-derived macrophage (BMDM) from WT and p38γ/δKIKO mice was exposed to LPS (100 ng/ml) for 0, 30, and 60 min and gene expression analysed by RNA-sequencing. Bar plot showing the number …
Excel files of data (raw and processed) showed in panels A–E.
(A) Table showing Gene Ontology (GO) biological processes genes. Negatively regulated genes are highlighted in bold. (B) Bone marrow-derived macrophage (BMDM) from WT and p38γ/δKIKO mice was exposed …
Excel files of data showed in panel A.
Labelled (.pdf) and raw (.tif) images from the arrays showed in panel B.
(A) Peritoneal macrophages from wild-type (WT), p38γ/δKO, and p38γ/δKIKO mice were exposed to lipopolysaccharide (LPS) (100 ng/ml) for 0 or 60 min and phosphorylated peptides identified. Heatmap …
Excel file of data showed in panels A–D and in Figure 4—figure supplement 1 panel C.
(A) Peritoneal macrophages from wild-type (WT), p38γ/δKO, and p38γ/δKIKO mice were exposed to lipopolysaccharide (LPS) (100 ng/ml) for the indicated times. Cell lysates (30 μg) were immunoblotted …
Labelled (.pdf) and raw (.jpeg) western blot images showed in panel A.
(A) Liquid chromatography–mass spectrometry (LC–MS)/MS analysis of the m/z ion of the phosphorylated MEF2D peptide (amino acids and phosphopeptide sequences are indicated, where pS (red) indicates …
Excel file of data showed in Figure 4—figure supplement 2.
(A) Bone marrow-derived macrophage (BMDM) from wild-type (WT) and p38γ/δKIKO mice was exposed to lipopolysaccharide (LPS) (100 ng/ml) for the indicated times. Relative mRNA expression of Jun, Cd14, …
Labelled (.ppt) western blot images showed in panel B.
Labelled (.ppt) western blot images showed in panel C.
Labelled (.ppt) western blot images showed in panel D.
Graph shows % survival at the indicated times. (Right panel) 30 μg of protein lysates from WT and LysMCre-p38γ/δ-/- peritoneal macrophage were immunoblotted with the indicated antibodies to total …
Human GST-MEF2D was incubated in a phosphorylation reaction mix containing Mg-ATP in the absence (control) or the presence of p38α or p38δ, and subjected to sodium dodecyl sulfate–polyacrylamide gel …
Protein phospho-site (Q14814) | Phosphopeptide | Control | p38α | p38δ |
---|---|---|---|---|
S98 | KGFN | GCD PEPDGEDSLEQSPLLEDK+ | +++ | - |
KGFNGCD | PEPDGEDSLEQSPLLEDK||||
GFNGCD | PEPDGEDSLEQSPLLEDK||||
GFNGCD | PEPDGEDSLEQSPLLEDKYR||||
S121 | RA | EELDGLFR+ | + | +++ |
S180 | LL | PQQPALQR- | +++ | +++ |
S192 | NSV | PGLPQR- | + | - |
S231 | A | PGLLPVANGNSLNK- | + | +++ |
A | PGLLPVAN GNSLNK||||
A | PGLLPVAN GN SLNK||||
S251 | VIPAK | PPPPTHSTQLGAPSR- | +++ | +++ |
PPPPTHSTQLGAPSR | ||||
S275 | VIT | QAGK- | - | +++ |
S444 | SEPV | PSRER- | - | +++ |
S450 | ER | PAPPPPAVFPAAR- | + | +++ |
S472 | PEPGDGLS | PAGGSYETGDR- | - | +++ |
Antibody | Name | Provider | Method | Dilution |
---|---|---|---|---|
P-ERK1/2 | Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody | Cell Signaling | WB | 1/2000 |
ERK1/2 | p44/42 MAPK (Erk1/2) Antibody | Cell Signaling | WB | 1/1000 |
TPL2 | Cot (M-20) | Santa Cruz Biotechnology, Inc | WB | 1/1000 |
ABIN2 | ABIN2 antibody | S.C. Ley laboratory | WB | 2 µg/ml |
P-p38 | Phospho-p38 MAP Kinase (Thr180/Tyr182) Antibody | Cell Signaling | WB | 1/1000 |
p38α | p38α (C-20) | Santa Cruz Biotechnology, Inc | WB | 1/1000 |
p38γ | p38γ (GST-SAPK3) S524A 1st Bleed | DSTT* | WB | 1/1000 |
p38δ | p38δ (GST-SAPK4) S526A 3rd Bleed | DSTT* | WB | 1/1000 |
P-JNK1/2 | Rabbit (polyclonal) Anti-JNK1&2 [pTpY183/185] Phosphospecific Antibody, Unconjugated | Biosource | WB | 1/1000 |
JNK1/2 | SAPK/JNK | Cell Signaling | WB | 1/1000 |
P-p105 | Phospho-NF-κB p105 (Ser933) (18E6) Rabbit mAb | Cell Signaling | WB | 1/1000 |
p105 | NF-κB1 p105 Antibody | Cell Signaling | WB | 1/1000 |
hDlg | hDlg (SAP97) antibody | DSTT* | WB | 1/1000 |
P-hDlg (S158) | Phospho-hDlg (SAP97) Ser158 | DSTT* | WB | 1/1000 |
hDlg | hDlg (SAP97) antibody | DSTT* | IP | 1 μg/IP |
p38γ | p38γ (SAPK3 C-Terminal) [KPPRNLGARVPKETA] | DSTT* | IP | 2 μg/IP |
CD45 | CD45 Monoclonal Antibody (30-F11), APC #17-0451-82 | eBioscience | FAC | 1/100 |
F4/80 | F4/80 Monoclonal Antibody (BM8) FITC #11-4801-85 | eBioscience | FAC | 1/100 |
Ly6G | PE Rat Anti-Mouse Ly-6G Clone 1A8 (RUO) #551461 | BD Bioscience | FAC | 1/50 |
DSTT (Dundee): Division of Signal Transduction Therapy; University of Dundee (Dundee, UK).
Gene | Forward (5′–3′) | Reverse (5′–3′) |
---|---|---|
Tnfa | TTGAGATCCATGCCGTTG | CTGTAGCCCACGTCGTAGC |
Il1b | TGGTGTGTGACGTTCCCATT | CAGCACGAGGCTTTTTTGTTG |
Il6 | GAGGATACCACTCCCAACAGACC | AAGTGCATCATCGTTGTTCATACA |
Ifnb1 | TCAGAATGAGTGGTGGTTGC | GACCTTTCAAATGCAGTAGATTCA |
Nos2 | CAGCTGGGCTGTACAAACCTT | CATTGGAAGTGAAGCGTTTCG |
Il12a | CCACCCTTGCCCTCCTAAA | GGCAGCTCCCTCTTGTTGTG |
Ccl5 | ATATGGCTCGGACACCACTC | TTCTTCGAGTGACAAACACG |
Cxcl9 | TGCACGATGCTCCTGCA | AGGTCTTTGAGGGATTTGTAGTGG |
Cxcl10 | GGATCCCTCTCGCAAGGA | ATCGTGGCAATGATCTCAACA |
Cxcl1 | CCTTGACCCTGAAGCTCCCT | CGGTGCCATCAGAGCAGTCT |
Ccl2 | TCTGGGCCTGCTGTTCACA | TTGGGATCATCTTGCTGGTG |
Cxcl2 | CCTGGTTCAGAAAATCATCCA | CTTCCGTTGAGGGACAGC |
Jun | GCACATCACCACTACACCGA | GGGAAGCGTGTTCTGGCTTAT |
Cd14 | CATTTGCATCCTCCTGGTTTCTGA | GAGTGGTTTTCCCCTTCCGTGTG |
Hdac7 | CGCAGCCAGTGTGAGTGTCT | AGTGGGTTCGTGCCGTAGAG |
Actb | AAGGAGATTACTTGCTCTGGCTCCTA | ACTCATCGTACTCCTGCTTGCTGAT |