Where to Stay/Visit in Tuscany (4 nights)

There are lots of Tuscany rental car discussions on the forum that might be help if you search.

With staying at an agriturismo and needing a car, your options are limited if you don't rent in Florence. You could take the train to Chiusi, I believe there are some rental car options there. Then you have a shorter drive, but you are driving back part of the way you came. The rental car locations outside Florence will have more restricted open hours and options for cars so options will also depend on what days of the week you are picking up/returning. But Chiusi also makes it easy to drop off there and take the train to Rome.

We stayed in Montalcino and rented from Florence Europcar (with pick up at Santa Maria Novella train station). We unfortunately left Florence at 5pm rush hour so there was a decent amount of traffic, but I think had it been earlier in the day it would be pretty easy to exit from the city center heading south or east/south. You get out of the city pretty quickly. This could give you the option to stop in Siena, if you have a car with a regular trunk so your luggage is hidden.

You really can't go wrong with any of these options. Personally I would pick Siena or Orvieto over Volterra and Sam Gimignano given where you are staying (i.e. furthest distance), and that they are more unique. V & SG would be more similar to what you'll get from Pienza, Montepulciano, and Montalcino.

Siena is probably my favorite of the whole group - I love the Piazza del Campo and walking around the various contrade/neighborhoods. Orvieto wasn't on my radar (friend suggested going) and I really enjoyed the tour through the old underground part of the city and learning about the history there.

We did a wonderful Road Scholar tour that has us five nights in Siena (used it as a base for Tuscany) and four nights in Spoleto as a base for Umbria.
I recommend Siena, since it is a great place to visit. I can't imagine staying in San Gimignano, I imagine the hotel rates there are $$$$$$.

We did Orvieto when visiting Umbria and loved it.

You might want to stay 3 nights in Venice, even though you arrive early, you may be jet lagged.
3 nights Florence, you can do an easy day trip to Siena via bus. Agree with going to Chiusi to get the rental car, stay around Pienza/Montepulciano, return car in Chiusi, and train to Rome.
Orvieto and Assisi are in Umbria BTW, and Orvieto has a rental car place by the train station, also.

I do like your itinerary as is. While Venice could use another night I wouldn’t want to cut from anywhere in your plan to accomplish that. (You will just have to plan to return)

You could pick up car in Chiusi but that will require an almost 2 hr train and getting there before the agency closes (Mid day, noon on Sat, closed Sunday). Your options will also be limited- automatic possibly not available, only Budget/Avis there.
You could drop the car there though and take train to Rome.

If you are worried about pick up in Florence you can pick up at Florence airport- take a taxi - right to the rental agency location- about 22 euros. It’s on the wrong side of where you are headed but overall will get you to Val D’Orcia quicker and you can take the Chianti road drive which is lovely.
If you want to drop in Chiusi rent from Budget/Avis- see what autoeurope.com offers.
Or you can drop in Orvieto if you rent from Hertz and take train to Rome from there (only agency in Orvieto).

All drivers must have an IDP- required by law. Make sure you do your homework about driving in Italy- ZTLs, parking, speed cameras, etc. Driving in the Tuscany region is easy and enjoyable. If you plan to do any wine tasting while out and about you will need a designated driver. We choose to just taste in town or at meals to avoid this issue.

Day trips
Definitely do:
Pienza, Montepulciano, Monticchiello, San Quirico/Bagno Vignoni.
You will not be driving IN to any of these towns but parking outside the walls in lots. All are pretty easy.

For a longer day trip— Siena or Assisi.
Siena really needs more than a day.
Assisi is a nice drive thru Umbria for a change of pace. Could visit Spello as well.
I would say play this by ear- you may just want a do nothing day after Venice/Florence and before Rome. Nothing wrong with that!

Do you already have lodging booked?

@ChristineH
I have booked accommodations for Venice, Florence, and Rome so far. Still finalizing the stay in Val D'Orcia.
And yes, playing it by ear and giving ourselves a nothing day was something I have thought about as well. After all, we don't know how we will feel mid-way through our trip. This is the biggest trip we've had together, which is why I like the idea of a Tuscan/Umbrian escape between Florence and Rome where we don't have to stress over an itinerary. It's nice to know there's nothing wrong with that! Thank you.

Based on the comments, I will look into a car rental from Florence. Seems like it could save precious time.

For your Pienza/ Val d'Orcia stay look into www.toscanaadue.com and www.theisabellaexperience.com for possible options. We have had wonderful experiences with both. Toscana Due is Niccolò Moricciani and his now fiancé Leonie and Niccolò is Isabella's son. Happy wedding and honeymoon planning!

Gene thank you for those recommendations!

There are a million ways to see Italy and an almost unlimited number of ways to do it. In Tuscany/Umbria, I also like to have a car. On our last trip we rented one in Orvieto. The Agra Turismo we stayed in was just outside of Civita di Bagnoregio. Our view was of the hilltop city. It was wonderful. From there we explored many of the places you have an interest in but we also visited a place that seems to get overlooked, Tarquinia.
Sienna is worth visiting. I have been there a number of times. On this trip we went there by bus from Florence. I just spoke to my sister about her visit there. She and her husband went by train and stayed there overnight. She recommended that. She is my younger sister so there is no way I can listen to her recommendation.
We returned the car in Orvieto and continued on to Rome by train.
I would recommend Tarquinia, Orvieto and Civita and Sienna. Florence is admittedly not my favorite place so with that for perspective, draw your conclusions about my advice.
If you are interested in the Agre Turismo we stayed at, I can PM you details. The owner was a wonderful person. We plan to eventually return there.