Clear the field Assignee when a sub-task has been assigned to the same user for the second time.
You can alternatively set an assignee automatically based on the members of a group.
Create a new rule and name it appropriately.
Providing a description will help you to identify what the rule does but this step is optional.
Field changed event
Add a Trigger → Field changed event
Choose Any value
Add a Condition → Boolean condition
Use the following expression:
Update field action
Add an Action → Update field action
Choose your custom Assignee
Choose No value - clear field