Studies

Macroautophagy can regulate cell signalling and tumorigenesis via elusive molecular mechanisms. We establish a RAS mutant cancer cell model where the autophagy gene ATG5 is dispensable in A549 cells in vitro, yet promotes tumorigenesis in mice. ATG5 represses transcriptional activation by the TGFβ-SMAD gene regulatory pathway. However, autophagy does not terminate cytosolic signal transduction by TGFβ. Instead, we use proteomics to identify selective degradation of the signalling scaffold TRAF3. ...

The human lysosomal polypeptide ABC transporter TAPL (ABC subfamily B member 9, ABCB9) transports 6–59 amino-acids-long polypeptides from the cytosol into lysosomes. The subcellular localization of TAPL depends solely on its N-terminal transmembrane domain TMD0, which lacks conventional targeting sequences. However, the intracellular route and the molecular mechanisms that control TAPL localization remain unclear. Here, we delineated the route of TAPL to lysosomes and investigated the determinants ...

HEK293T cells stably expressing N-terminally HA-tagged BAG3 were employed to screen for novel BAG3 interacting proteins.

SILAC-based proximity proteomics of UAPEX2-tagged UBE2QL1 in differential LLOMe-treated HeLa cells

We selected GABARAPL2endoHA cells for a proof-of-principle immunoprecipitation (IP) followed by mass spectrometric (MS) analysis to identify new binding partner candidates of a hATG8 family member at endogenous levels. To distinguish between candidates that bind preferentially to PE-conjugated versus unconjugated GABARAPL2 we treated stable isotope labeling with amino acids in cell culture (SILAC)-labeled GABARAPL2endoHA cells with Torin1 and BafA1 (light) or ATG7 inhibitor (heavy).

Proximity proteomics of GAL9 upon lysosomal damage by LLOMe or GPN. Proximity labeling was performed in SILAC labelled HEK293T cells stably expressing APEX2-GAL9.

Identification of autophagy substrates by directing APEX2 to autophagosomes and immune isolation of lysosomes.