Development of a Signal-integrating Reporter to Monitor\nMitochondria–ER Contacts

Mitochondria–endoplasmic reticulum\ncontact sites (MERCS)\nserve as hotspots for important cellular processes, including calcium\nhomeostasis, phospholipid homeostasis, mitochondria dynamics, and\nmitochondrial quality control. MERCS reporters based on complementation\nof green fluorescent proteins (GFP) fragments have been designed to\nvisualize MERCS in real-time, but we find that they do not accurately\nrespond to changes in MERCS content. Here, we utilize split LacZ complementing\nfragments to develop the first MERCS reporter system (termed SpLacZ-MERCS)\nthat continuously integrates the MERCS information within a cell and\ngenerates a fluorescent output. Our system exhibits good organelle\ntargeting, no artifactual tethering, and effective, dynamic tracking\nof the MERCS level in single cells. The SpLacZ-MERCS reporter was\nvalidated by drug treatments and genetic perturbations known to affect\nmitochondria–ER contacts. The signal-integrating nature of\nSpLacZ-MERCS may enable systematic identification of genes and drugs\nthat regulate mitochondria–ER interactions. Our successful\napplication of the split LacZ complementation strategy to study MERCS\nmay be extended to study other forms of interorganellar crosstalk.