The fret inlays (assuming they're vertical - those are the easiest to distinguish with) will tell you were the actual fret would be installed on the same neck. Since different types of guitars have different scale lengths, these are important to show you were the note sits on the fretboard.
Technically it wouldn't matter what you tuned your bass to, as long as it was probably intonated, those fret markers would should you were the next western musical note would lie on that particular fretboard/bass.
You do want to play by ear, yes. But you also want to be somewhat academic about it since you're starting out on fretless bass.
Check out this vector demonstration of what's going on:
You want to "fret" the note on the fret marker as much as possible. Eventually you'll get a feel for it and your fretless playing will sound as in-tune as possible. Then, utilize that ear and squeeze notes where they should go, using the other instruments in the band for reference points (But make sure they're tuned too!)
Caveat: you may not know it, but you're probably not putting your fingers in the exact same place every time you play your bass. Your frets assist you when that happens. So don't rely on "always being in the center of the fret". And, in reality, the closer you can get to the fret (on fretted instrument), the less pressure you require to push that string down and get a good note out. And therefore, the closer your finger gets used to being where the fret is allowing it to learn that same location for fretless applications.