Hey @tamberg, great question! You absolutely can plug USB cables into both the AF and the ESP32 at the same time. I do this all the time during development for exactly the reason you describe: to connect directly to the Notecard over USB serial and push firmware updates to the ESP32. Both devices will draw their power directly from those ports, but both are connected to a common ground, so won’t interfere when simultaneously connected.
As for the JST connectors, we recommend powering your project through the JST port on the Notecarrier-AF and not from the ESP32 or another Feather that includes a LiPo. You’re not at risk of ruining either device by powering through the Feather JST in terms of what we have tested, but since the Notecarrier includes additional power circuitry and reverse polarity protection, you’re better off going through the JST on the Notecarrier itself.