Rental Car vs. Train

Grant, first, you don't need a rental car in Venice. The whole place is on water. Thus, I suspect that you would rent a car from the larger car rental facility at the Tronchetto when departing. Caution - do not go 1MPH over the speed limit on the causeway departing the islands of Venice. Second, tourist cars are not permitted in any of the five CT towns. You are permitted to enter to get to the parking lots, which are full in the summer, or to your hotel should it provide parking. Your rental car is useless in the CT.

Tuscany and Umbria are the best areas to have a rental car. So much to see and the train/bus systems there are limited. You can rent your car in the Siena area, drive all around, and when you are ready for Rome, drive south to Orvieto to turn in your rental car. Orvieto is a very popular area to pick up and drop off rental cars. The Hertz office is about 100 yards from the train station. So is a fuel station to fill up your fuel tank (usually diesel) before turning it in.

Finally, off to Rome you go on the train. You don't want to drive into Rome looking for a place to turn in your rental car.

You could rent a car in LaSpezia the day you leave CT. There's a Hertz rental a quick cab ride from the train station, and you only have 2 turns to get on the autostrada. We did this, then drove to Voltera and San Gimignano on the way our stay in Montalcino. Then we dropped the car in Orvieto, and took the train to Rome the following day.

I would train from Venice to CT since as noted above cars are useless in each city. I did the trip and it's about a 5 hour (to 6 hour depending on which train you select) trip with at least 2 connections. (I went from Venice - Florence, Florence - La Spezia, La Spezia - Vernazza). We left Venice early in the AM to get a jump on the day.

For Tuscany how many days are you there and are you staying in Siena the whole time for your hotel or are you going to say stay in Volterra a night, another village another night? A car is your best bet to explore the tuscany towns and countryside. Just note most cities you can't drive in, and also note the ZTL's around Siena. Siena has some bus connections to the bigger hilltowns that work, but if you want to explore the vineyards and real small quaint cities, grab a rental.

"A couple of days in each city" doesn't give you any time to do daytrips. In your shoes, I would do trains and not deal with the hassle of renting a car and learning about driving in a foreign country. And on a train you can move around or take a nap or use the bathroom or read. Not possible if you're driving. (Plus you might meet interesting people on the train.)

Your itinerary doesn't require a car and in some cases it would be a hassle. It's easy to train from Venice to the Cinque Terre and then to Siena. Siena could be a day base to reach Florence. Or other hilltowns. You could reach Orvieto, Pisa and Assisi from Siena on the train. Or you could bus from Siena to San Gimignano, Volterra, Montepulciano and more. Then train from Siena to Rome. Usually for 2 people the cost of the car isn't worth it. Plus, in Italy there is mandatory theft insurance, tolls on the highways, parking fees, high fuel costs and hilltowns and some cities impose driving restrictions. Its very easy to end up with multiple driving violations without knowing it. The train might be the way to go and more relaxing.

Forget the car. you have no time to stray around in your itinerary.
Go from Venice to Tuscany, THEN To Cinque Terre, finally from Cinque Terre to Rome.

All - Thanks for all the great feedback! We appreciate the advice and can't wait to explore Italy.
Cheers!
Grant

You should strongly consider Roberto's suggestion for reordering your itinerary. He knows what he's talking about.