Classic ASP suddenly giving me permissions (401.3) error

Background: I support a classic ASP environment. I have a development setup locally on my machine as part of that support. I am running IIS7.

To access my environment, I use "http://localhost:99999/" (port # faked for privacy purposes). I have not had a problem with this -- until today.

I built an application that is intended to run in this environment. The app is ASP.NET v.4.0 (for sake of example, I'll say it's called "http://localhost:99999/DotNetApp/"). Of course, in order to run this, I had to set up my environment application pool to support it (which it now does).

Problem: after configuring this, I am now getting:

Server Error in '/' Application.

Access is denied.
Description: An error occurred while accessing the resources required to serve this request. You might not have permission to view the requested resources.

Error message 401.3: You do not have permission to view this directory or page using the credentials you supplied (access denied due to Access Control Lists). Ask the Web server's administrator to give you access to '[filepath]\SourceFiles'.

I did NOT get this before I set my IIS configuration. Why am I getting this now?

Note: this ONLY happens with "http://localhost:99999/"; this does NOT happen if I try "http://localhost:99999/default.asp" (the page comes up with no problem).

Anyone have any insight?

Thanks in advance . . .

Edit: Additional symptoms: I tried playing with the application pool settings. The problem goes away if I change the .NET Framework version to either 2.0.50727 or to "No Managed Code." Changing it to 4.0.31319 breaks it. Of course, if I use any of the other two versions, my ASP.NET app won't run.

Edit #2: This problem occurs on ALL links that go to a folder or directory (e.g. "http://localhost:99999/somefolder/"), not just the web root.

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