diff --git a/changelog.d/9473.bugfix b/changelog.d/9473.bugfix
new file mode 100644
index 0000000000..71fb487cf2
--- /dev/null
+++ b/changelog.d/9473.bugfix
@@ -0,0 +1 @@
+Fix long-standing bug when generating thumbnails for some images with transparency: `TypeError: cannot unpack non-iterable int object`.
diff --git a/synapse/rest/media/v1/thumbnailer.py b/synapse/rest/media/v1/thumbnailer.py
index 07903e4017..988f52c78f 100644
--- a/synapse/rest/media/v1/thumbnailer.py
+++ b/synapse/rest/media/v1/thumbnailer.py
@@ -96,9 +96,14 @@ class Thumbnailer:
     def _resize(self, width: int, height: int) -> Image:
         # 1-bit or 8-bit color palette images need converting to RGB
         # otherwise they will be scaled using nearest neighbour which
-        # looks awful
-        if self.image.mode in ["1", "P"]:
-            self.image = self.image.convert("RGB")
+        # looks awful.
+        #
+        # If the image has transparency, use RGBA instead.
+        if self.image.mode in ["1", "L", "P"]:
+            mode = "RGB"
+            if self.image.info.get("transparency", None) is not None:
+                mode = "RGBA"
+            self.image = self.image.convert(mode)
         return self.image.resize((width, height), Image.ANTIALIAS)
 
     def scale(self, width: int, height: int, output_type: str) -> BytesIO:
diff --git a/tests/rest/media/v1/test_media_storage.py b/tests/rest/media/v1/test_media_storage.py
index 36d1e6bc4a..9f77125fd4 100644
--- a/tests/rest/media/v1/test_media_storage.py
+++ b/tests/rest/media/v1/test_media_storage.py
@@ -105,7 +105,7 @@ class MediaStorageTests(unittest.HomeserverTestCase):
         self.assertEqual(test_body, body)
 
 
-@attr.s
+@attr.s(slots=True, frozen=True)
 class _TestImage:
     """An image for testing thumbnailing with the expected results
 
@@ -117,13 +117,15 @@ class _TestImage:
             test should just check for success.
         expected_scaled: The expected bytes from scaled thumbnailing, or None if
             test should just check for a valid image returned.
+        expected_found: True if the file should exist on the server, or False if
+            a 404 is expected.
     """
 
     data = attr.ib(type=bytes)
     content_type = attr.ib(type=bytes)
     extension = attr.ib(type=bytes)
-    expected_cropped = attr.ib(type=Optional[bytes])
-    expected_scaled = attr.ib(type=Optional[bytes])
+    expected_cropped = attr.ib(type=Optional[bytes], default=None)
+    expected_scaled = attr.ib(type=Optional[bytes], default=None)
     expected_found = attr.ib(default=True, type=bool)
 
 
@@ -153,6 +155,21 @@ class _TestImage:
                 ),
             ),
         ),
+        # small png with transparency.
+        (
+            _TestImage(
+                unhexlify(
+                    b"89504e470d0a1a0a0000000d49484452000000010000000101000"
+                    b"00000376ef9240000000274524e5300010194fdae0000000a4944"
+                    b"4154789c636800000082008177cd72b60000000049454e44ae426"
+                    b"082"
+                ),
+                b"image/png",
+                b".png",
+                # Note that we don't check the output since it varies across
+                # different versions of Pillow.
+            ),
+        ),
         # small lossless webp
         (
             _TestImage(
@@ -162,8 +179,6 @@ class _TestImage:
                 ),
                 b"image/webp",
                 b".webp",
-                None,
-                None,
             ),
         ),
         # an empty file
@@ -172,9 +187,7 @@ class _TestImage:
                 b"",
                 b"image/gif",
                 b".gif",
-                None,
-                None,
-                False,
+                expected_found=False,
             ),
         ),
     ],