diff --git a/django/db/models/sql/query.py b/django/db/models/sql/query.py
index a39430b28d..b524e8859f 100644
--- a/django/db/models/sql/query.py
+++ b/django/db/models/sql/query.py
@@ -9,7 +9,6 @@ all about the internals of models in order to get the information it needs.
 import copy
 import difflib
 import functools
-import inspect
 import sys
 import warnings
 from collections import Counter, namedtuple
@@ -2038,19 +2037,9 @@ class Query(BaseExpression):
         group_by = list(self.select)
         if self.annotation_select:
             for alias, annotation in self.annotation_select.items():
-                signature = inspect.signature(annotation.get_group_by_cols)
-                if 'alias' not in signature.parameters:
-                    annotation_class = annotation.__class__
-                    msg = (
-                        '`alias=None` must be added to the signature of '
-                        '%s.%s.get_group_by_cols().'
-                    ) % (annotation_class.__module__, annotation_class.__qualname__)
-                    warnings.warn(msg, category=RemovedInDjango40Warning)
-                    group_by_cols = annotation.get_group_by_cols()
-                else:
-                    if not allow_aliases or alias in column_names:
-                        alias = None
-                    group_by_cols = annotation.get_group_by_cols(alias=alias)
+                if not allow_aliases or alias in column_names:
+                    alias = None
+                group_by_cols = annotation.get_group_by_cols(alias=alias)
                 group_by.extend(group_by_cols)
         self.group_by = tuple(group_by)
 
diff --git a/docs/releases/4.0.txt b/docs/releases/4.0.txt
index 43e94a790b..339147201d 100644
--- a/docs/releases/4.0.txt
+++ b/docs/releases/4.0.txt
@@ -257,6 +257,9 @@ to remove usage of these features.
 * ``django.views.i18n.set_language()`` doesn't set the user language in
   ``request.session`` (key ``_language``).
 
+* ``alias=None`` is required in the signature of
+  ``django.db.models.Expression.get_group_by_cols()`` subclasses.
+
 See :ref:`deprecated-features-3.1` for details on these changes, including how
 to remove usage of these features.
 
diff --git a/tests/expressions/test_deprecation.py b/tests/expressions/test_deprecation.py
deleted file mode 100644
index cdb1e43af6..0000000000
--- a/tests/expressions/test_deprecation.py
+++ /dev/null
@@ -1,24 +0,0 @@
-from django.db.models import Count, Func
-from django.test import SimpleTestCase
-from django.utils.deprecation import RemovedInDjango40Warning
-
-from .models import Employee
-
-
-class MissingAliasFunc(Func):
-    template = '1'
-
-    def get_group_by_cols(self):
-        return []
-
-
-class GetGroupByColsTest(SimpleTestCase):
-    def test_missing_alias(self):
-        msg = (
-            '`alias=None` must be added to the signature of '
-            'expressions.test_deprecation.MissingAliasFunc.get_group_by_cols().'
-        )
-        with self.assertRaisesMessage(RemovedInDjango40Warning, msg):
-            Employee.objects.values(
-                one=MissingAliasFunc(),
-            ).annotate(cnt=Count('company_ceo_set'))