Phospho jnk1 2

The following primary Abs were used in this study: Aggrecan (6-B-4) (Abcam, Cambridge, MA, USA), ARGXX (BC-3) (Abcam), phospho-JNK1/2 (Thermo Fisher Scientific, Waltham, MA, USA) β-actin (Cell Signaling Technology, Danvers, MA, USA), FLAG-M2 (MilliporeSigma, Burlington, MA, USA), acetylated α-tubulin (6-11B-1) (MilliporeSigma), early endosome antigen 1 (EEA-1; Abcam), LRP-1 β-light chain (Abcam), LRP-1 α-heavy chain (8G1) (Abcam), a disintegrin and metalloproteinase (ADAM)-17 (Abcam), MMP-13 (Santa Cruz Biotechnology, Dallas, TX, USA), MMP14 (anti-catalytic domain EP1264Y) (Abcam), mouse monoclonal anti–MT1-MMP hemopexin domain 222-1D8 (a gift from Yoshi Itoh, Kennedy Institute) generated as previously described (41 (link)), and arl13b (ProteinTech, Rosemont, IL, USA). Anti-AGEG was a kind gift from Hideaki Nagase (University of Oxford). Purified aggrecan from bovine cartilage came from MilliporeSigma. Recombinant human C-terminal His-tagged human receptor-associated protein (RAP) was produced in Escherichia coli using a pET3a-based expression vector and purified as described previously (13 (link)). The domain deletion mutant, ADAMTS-5 (TS5-3)-flag, is described in Gendron et al. (42 (link)), and recombinant MMP-13 is described in Yamamoto et al. (43 (link)). Recombinant IL-1β was purchased from PeproTech (London, United Kingdom).

Primary antibodies used in this study are as follows: β-actin (Novus Biologicals, 8H10D10 1:5000), acetylated α-tubulin (clone 6-11B-1, Sigma-Aldrich T7541 1:2000), arl13b (Proteintech, 17711-1-AP 1:1000), IFT88 (Proteintech, 13967-1-AP 1:500), IFT88 (Fig. 5B) (Santa Cruz Biotchnology, sc-84318 1:250), KIF3a (Abcam, ab11259 1:500), COX2 (Cell Signaling Technology, D5H5/12282 1:500), iNOS (Cell Signaling Technology D6B6S/13120 1:500), NFκB p65 (Abcam, ab7970 1:500), IKB (Abcam, ab32518 1:250), phospho-JNK1/2 (Thermo Fisher Scientific, D12H7L17) 1:500, phospho-ERK1/2 (Cell Signaling Technology, 9101 1:500). JLP/SPAG9 (D72F4) rabbit monoclonal antibody (Proteintech 5519 1:50), and γ-tubulin clone GTU-88 mouse monoclonal (Sigma, T6657 1:1000). Alexa Fluor-conjugated secondary antibodies (Invitrogen) and DAPI (1:5000, counterstain, also Invitrogen) were used for immunofluorescence studies.