diff --git a/.changeset/eleven-countries-hang.md b/.changeset/eleven-countries-hang.md
new file mode 100644
index 00000000000..fc25ba2c85a
--- /dev/null
+++ b/.changeset/eleven-countries-hang.md
@@ -0,0 +1,5 @@
+---
+'@primer/react': patch
+---
+
+Add support for dynamic aria-expanded support on TreeView.Item
diff --git a/src/TreeView/TreeView.stories.tsx b/src/TreeView/TreeView.stories.tsx
index e186675e1b0..ff4092d5af3 100644
--- a/src/TreeView/TreeView.stories.tsx
+++ b/src/TreeView/TreeView.stories.tsx
@@ -442,6 +442,9 @@ export const AsyncWithCount: Story = args => {
           </TreeView.Item>
           <TreeView.LinkItem href="#src">
             <TreeView.LeadingVisual>
+              <TreeView.LeadingVisual>
+                <TreeView.DirectoryIcon />
+              </TreeView.LeadingVisual>
               <TreeView.DirectoryIcon />
             </TreeView.LeadingVisual>
             src
@@ -622,4 +625,48 @@ export const StressTest: Story = () => {
   )
 }
 
+export const EmptyDirectory: Story = () => {
+  const [state, setState] = React.useState<SubTreeState>('loading')
+  const timeoutId = React.useRef<ReturnType<typeof setTimeout> | null>(null)
+
+  React.useEffect(() => {
+    return () => {
+      if (timeoutId.current) {
+        clearTimeout(timeoutId.current)
+        timeoutId.current = null
+      }
+    }
+  }, [])
+
+  return (
+    <Box sx={{p: 3, maxWidth: 400}}>
+      <TreeView aria-label="Test">
+        <TreeView.Item
+          onExpandedChange={expanded => {
+            if (expanded) {
+              timeoutId.current = setTimeout(() => {
+                setState('done')
+                timeoutId.current = null
+              }, 2000)
+            }
+          }}
+        >
+          <TreeView.LeadingVisual>
+            <TreeView.DirectoryIcon />
+          </TreeView.LeadingVisual>
+          src
+          <TreeView.SubTree state={state} />
+        </TreeView.Item>
+        <TreeView.Item>
+          <TreeView.LeadingVisual>
+            <TreeView.DirectoryIcon />
+          </TreeView.LeadingVisual>
+          .github
+          <TreeView.SubTree />
+        </TreeView.Item>
+      </TreeView>
+    </Box>
+  )
+}
+
 export default meta
diff --git a/src/TreeView/TreeView.test.tsx b/src/TreeView/TreeView.test.tsx
index c8b4c2d143b..4072d8955a2 100644
--- a/src/TreeView/TreeView.test.tsx
+++ b/src/TreeView/TreeView.test.tsx
@@ -1,8 +1,12 @@
 import {fireEvent, render, waitFor} from '@testing-library/react'
+import userEvent from '@testing-library/user-event'
 import React from 'react'
+import {act} from 'react-dom/test-utils'
 import {ThemeProvider} from '../ThemeProvider'
 import {SubTreeState, TreeView} from './TreeView'
 
+jest.useFakeTimers()
+
 // TODO: Move this function into a shared location
 function renderWithTheme(
   ui: Parameters<typeof render>[0],
@@ -155,6 +159,40 @@ describe('Markup', () => {
     // space
     expect(noDescription).toHaveAccessibleDescription(' ')
   })
+
+  it('should include `aria-expanded` when a SubTree contains content', async () => {
+    const user = userEvent.setup({
+      advanceTimers: jest.advanceTimersByTime
+    })
+    const {getByLabelText, getByText} = renderWithTheme(
+      <TreeView aria-label="Test tree">
+        <TreeView.Item>
+          Item 1
+          <TreeView.SubTree>
+            <TreeView.Item>Item 1.a</TreeView.Item>
+            <TreeView.Item>Item 1.b</TreeView.Item>
+            <TreeView.Item>Item 1.c</TreeView.Item>
+          </TreeView.SubTree>
+        </TreeView.Item>
+        <TreeView.Item>
+          Item 2
+          <TreeView.SubTree />
+        </TreeView.Item>
+      </TreeView>
+    )
+
+    let treeitem = getByLabelText(/Item 1/)
+    expect(treeitem).toHaveAttribute('aria-expanded', 'false')
+
+    await user.click(getByText(/Item 1/))
+    expect(treeitem).toHaveAttribute('aria-expanded', 'true')
+
+    treeitem = getByLabelText(/Item 2/)
+    expect(treeitem).not.toHaveAttribute('aria-expanded')
+
+    await user.click(getByText(/Item 2/))
+    expect(treeitem).not.toHaveAttribute('aria-expanded')
+  })
 })
 
 describe('Keyboard interactions', () => {
@@ -940,6 +978,10 @@ describe('Asyncronous loading', () => {
     // Click done button to mimic the completion of async loading
     fireEvent.click(doneButton)
 
+    act(() => {
+      jest.runAllTimers()
+    })
+
     // Live region should be updated
     expect(liveRegion).toHaveTextContent('Parent content loaded')
   })
@@ -983,13 +1025,15 @@ describe('Asyncronous loading', () => {
     // Wait for async loading to complete
     const firstChild = await findByRole('treeitem', {name: 'Child 1'})
 
-    setTimeout(() => {
-      // First child should be focused
-      expect(firstChild).toHaveFocus()
+    act(() => {
+      jest.runAllTimers()
     })
+
+    // First child should be focused
+    expect(firstChild).toHaveFocus()
   })
 
-  it.only('moves focus to parent item after closing error dialog', async () => {
+  it('moves focus to parent item after closing error dialog', async () => {
     function TestTree() {
       const [error, setError] = React.useState('Test error')
 
@@ -1064,4 +1108,53 @@ describe('Asyncronous loading', () => {
     // Parent item should still be expanded
     expect(parentItem).toHaveAttribute('aria-expanded', 'true')
   })
+
+  it('should remove `aria-expanded` if no content is loaded in', async () => {
+    function Example() {
+      const [state, setState] = React.useState<SubTreeState>('loading')
+      const timeoutId = React.useRef<ReturnType<typeof setTimeout> | null>(null)
+
+      React.useEffect(() => {
+        return () => {
+          if (timeoutId.current) {
+            clearTimeout(timeoutId.current)
+            timeoutId.current = null
+          }
+        }
+      }, [])
+
+      return (
+        <TreeView aria-label="Test tree">
+          <TreeView.Item
+            onExpandedChange={expanded => {
+              if (expanded) {
+                timeoutId.current = setTimeout(() => {
+                  setState('done')
+                }, 1000)
+              }
+            }}
+          >
+            Item 1
+            <TreeView.SubTree state={state} />
+          </TreeView.Item>
+        </TreeView>
+      )
+    }
+    const {getByLabelText, getByText} = renderWithTheme(<Example />)
+    const user = userEvent.setup({
+      advanceTimers: jest.advanceTimersByTime
+    })
+
+    const treeitem = getByLabelText(/Item 1/)
+    expect(treeitem).toHaveAttribute('aria-expanded', 'false')
+
+    await user.click(getByText(/Item 1/))
+    expect(treeitem).toHaveAttribute('aria-expanded', 'true')
+
+    act(() => {
+      jest.runAllTimers()
+    })
+
+    expect(treeitem).not.toHaveAttribute('aria-expanded')
+  })
 })
diff --git a/src/TreeView/TreeView.tsx b/src/TreeView/TreeView.tsx
index 3ea37f1bb4a..25c4ff34aa1 100644
--- a/src/TreeView/TreeView.tsx
+++ b/src/TreeView/TreeView.tsx
@@ -35,6 +35,8 @@ const RootContext = React.createContext<{
 const ItemContext = React.createContext<{
   itemId: string
   level: number
+  isSubTreeEmpty: boolean
+  setIsSubTreeEmpty: React.Dispatch<React.SetStateAction<boolean>>
   isExpanded: boolean
   setIsExpanded: React.Dispatch<React.SetStateAction<boolean>>
   leadingVisualId: string
@@ -42,6 +44,8 @@ const ItemContext = React.createContext<{
 }>({
   itemId: '',
   level: 1,
+  isSubTreeEmpty: false,
+  setIsSubTreeEmpty: () => {},
   isExpanded: false,
   setIsExpanded: () => {},
   leadingVisualId: '',
@@ -135,6 +139,7 @@ const Item = React.forwardRef<HTMLElement, TreeViewItemProps>(
     })
     const {level} = React.useContext(ItemContext)
     const {hasSubTree, subTree, childrenWithoutSubTree} = useSubTree(children)
+    const [isSubTreeEmpty, setIsSubTreeEmpty] = React.useState(!hasSubTree)
 
     // Expand or collapse the subtree
     const toggle = React.useCallback(
@@ -189,6 +194,8 @@ const Item = React.forwardRef<HTMLElement, TreeViewItemProps>(
         value={{
           itemId,
           level: level + 1,
+          isSubTreeEmpty,
+          setIsSubTreeEmpty,
           isExpanded,
           setIsExpanded,
           leadingVisualId,
@@ -205,7 +212,7 @@ const Item = React.forwardRef<HTMLElement, TreeViewItemProps>(
           aria-labelledby={labelId}
           aria-describedby={`${leadingVisualId} ${trailingVisualId}`}
           aria-level={level}
-          aria-expanded={hasSubTree ? isExpanded : undefined}
+          aria-expanded={isSubTreeEmpty ? undefined : isExpanded}
           aria-current={isCurrentItem ? 'true' : undefined}
           onKeyDown={handleKeyDown}
           sx={{
@@ -328,7 +335,7 @@ const Item = React.forwardRef<HTMLElement, TreeViewItemProps>(
               </Slots>
             </Box>
           </Box>
-          {isExpanded ? subTree : null}
+          {subTree}
         </Box>
       </ItemContext.Provider>
     )
@@ -409,11 +416,26 @@ export type TreeViewSubTreeProps = {
 
 const SubTree: React.FC<TreeViewSubTreeProps> = ({count, state, children}) => {
   const {announceUpdate} = React.useContext(RootContext)
-  const {itemId, isExpanded} = React.useContext(ItemContext)
+  const {itemId, isExpanded, isSubTreeEmpty, setIsSubTreeEmpty} = React.useContext(ItemContext)
   const [isLoadingItemVisible, setIsLoadingItemVisible] = React.useState(false)
   const {safeSetTimeout, safeClearTimeout} = useSafeTimeout()
   const timeoutId = React.useRef<number>(0)
   const loadingItemRef = React.useRef<HTMLElement>(null)
+  const ref = React.useRef<HTMLElement>(null)
+
+  React.useEffect(() => {
+    // If `state` is undefined, we're working in a synchronous context and need
+    // to detect if the sub-tree has content. If `state === 'done` then we're
+    // working in an asynchronous context and need to see if there is content
+    // that has been loaded in.
+    if (state === undefined || state === 'done') {
+      if (!isSubTreeEmpty && !children) {
+        setIsSubTreeEmpty(true)
+      } else if (isSubTreeEmpty && children) {
+        setIsSubTreeEmpty(false)
+      }
+    }
+  }, [state, isSubTreeEmpty, setIsSubTreeEmpty, children])
 
   // Announce when content has loaded
   React.useEffect(() => {
@@ -422,10 +444,18 @@ const SubTree: React.FC<TreeViewSubTreeProps> = ({count, state, children}) => {
 
       if (!parentItem) return
 
+      const {current: node} = ref
       const parentName = getAccessibleName(parentItem)
-      announceUpdate(`${parentName} content loaded`)
+
+      safeSetTimeout(() => {
+        if (node && node.childElementCount > 0) {
+          announceUpdate(`${parentName} content loaded`)
+        } else {
+          announceUpdate(`${parentName} is empty`)
+        }
+      })
     }
-  }, [state, itemId, announceUpdate])
+  }, [state, itemId, announceUpdate, safeSetTimeout])
 
   // Show loading indicator after a short delay
   React.useEffect(() => {
@@ -462,6 +492,10 @@ const SubTree: React.FC<TreeViewSubTreeProps> = ({count, state, children}) => {
     }
   }, [state, safeSetTimeout, safeClearTimeout, isLoadingItemVisible, itemId])
 
+  if (!isExpanded) {
+    return null
+  }
+
   return (
     <Box
       as="ul"
@@ -472,6 +506,8 @@ const SubTree: React.FC<TreeViewSubTreeProps> = ({count, state, children}) => {
         padding: 0,
         margin: 0
       }}
+      // @ts-ignore Box doesn't have type support for `ref` used in combination with `as`
+      ref={ref}
     >
       {isLoadingItemVisible ? <LoadingItem ref={loadingItemRef} count={count} /> : children}
     </Box>