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Summary: In mammalian cells, the Golgi exists in ribbon architecture—individual stacks laterally linked to each other by tubular structures. Golgi architecture changes dynamically to cater to cellular needs. Loss of architecture is linked with pathological conditions like cancer and neurodegeneration. Not much is known about the regulators of Golgi dynamics. Here, we demonstrate that CARP2 (caspase-8- and caspase-10-associated RING-containing protein 2), an endosomal ubiquitin ligase and a known regulator of cell migration, modulates Golgi dynamics. Epidermal growth factor (EGF) treatment modestly increases CARP2 protein and disperses Golgi. An exogenous supply of CARP2 also leads to Golgi dispersal. Conversely, Golgi remains intact in CARP2 knockout (KO) cells upon EGF treatment. CARP2 variants defective in either endosomal association or ligase activity are unable to affect Golgi dispersal. Importantly, CARP2 targets Golgin45 for ubiquitination and degradation in EGF-stimulated cells. Collectively, our findings unravel the existence of crosstalk between endosomal ubiquitin signaling and Golgi dynamics.

Caspases are responsible for the execution of programmed cell death (apoptosis) and must undergo proteolytic activation, in response to apoptotic stimuli, to function. The mechanism of initiator caspase activation has been generalized by the induced proximity model, which is thought to drive dimerization-mediated activation of caspases. The initiator caspase, caspase-9, exists predominantly as a monomer in solution. To examine the induced proximity model, we engineered a constitutively dimeric caspase-9 by relieving steric hindrance at the dimer interface. Crystal structure of the engineered caspase-9 closely resembles that of the wild-type (WT) caspase-9, including all relevant structural details and the asymmetric nature of two monomers. Compared to the WT caspase-9, this engineered dimer exhibits a higher level of catalytic activity in vitro and induces more efficient cell death when expressed. However, the catalytic activity of the dimeric caspase-9 is only a small fraction of that for the Apaf-1-activated caspase-9. Furthermore, in contrast to the WT caspase-9, the activity of the dimeric caspase-9 can no longer be significantly enhanced in an Apaf-1-dependent manner. These findings suggest that dimerization of caspase-9 may be qualitatively different from its activation by Apaf-1, and in conjunction with other evidence, posit an induced conformation model for the activation of initiator caspases.