When to use ultimates in Junkenstein's Revenge?
McCree is the character that will deal the most damage to "Bosses" in a short delay of time, so Ana boosting McCree to deal with bosses faster is a good strategy (Because yes, boosting McCree does increase the charge speed).
Everyone should try to focus as much as possible the grenadier since they are the only mob that will injure you.
In depth Analysis
McCree
I personally like having McCree top left to keep him safe when bosses spawn and far as much as possible from Roadhog. Since he have a good hitscan, he can deal with bots on the other side of the map. Don't forget to roll with him to reload faster once in a while. And use the flashbang to stop grenadiers for a second or to deal some extra damage to tires.
Make sure you are not the focus target of any boss before launching your ult. Junkrat's bombs (which does a whooping 120 damage on contact) combined with the slowing effect of your ult should lay you down pretty quickly. Roadhog would just hook and take you out of your ult before you can do anything.
Quick heads up, it might be tempting to wait until the very last second to ult because "you'll be able to oneshot them in just a fraction of a second more". Don't. Shoot it, don't be greedy. It happened to me too many times. Your teamates will finish them off. Which wouldn't happen if you didn't shoot at all.
Soldier 76
Soldier 76 is usually in the middle and uses his ult as frequently as he can to clear huge waves of bots. Grenades should be used to deal with grenadiers since it gets rid of 75% of their hp. Since his damage burst is next to inexistant, you should probably defend the gate hp over dealing damage to bosses (except for the grenade who can either deal with grenadier or do a bunch of damage to bosses)
Hanzo
Hanzo likes to be located on a bit higher ground to get good scatter arrows. So 2 good locations would be on the right ledge next to the defender door or on Junkenstein's platform before he spawns. Keep in mind that having him on Junkenstein's platform makes it so no one is really watching the right path. Sonic arrow can be used to show ennemies in blind spots of your team try to use it accordingly. Finally, your ult covers Junkenstein's platform if you launch it on the corner of the tower from the door. Don't be too shy to use it on something else than bosses as it covers an entire path of bots.
Ana
Ana should be able to roam freely with the positionning that I described, this way she can heal everyone easily. Sleep dart should be used to ease the grenadier flow (Sleeping one while the others are being cared of). Sleep dart is also of good use when being used after Reaper's Wraith Form. This way it guarranties McCree's ult to land. Just make sure to land it in the last 3-4 seconds of McCree's channel. Communication with him will ease the process also. Healing grenade is used for both healing and to get rid of tires that gets too near (since you should always be somewhat near the door). And her ult should be used for McCree as said previously or 76 if there's really too much bots coming your way. Shooting enemies with Ana should be only a 1 hit tag (except for grenadier). It makes enemies easier to destroy for your teamates and ensures that you have a good view of the whole battlefield to keep everyone topped off.