diff --git a/core/src/components.d.ts b/core/src/components.d.ts
index d78d21b79f..f5f6867991 100644
--- a/core/src/components.d.ts
+++ b/core/src/components.d.ts
@@ -2484,6 +2484,7 @@ export namespace Components {
           * The mode determines which platform styles to use.
          */
         "mode"?: "ios" | "md";
+        "setFocus": () => Promise<void>;
         /**
           * The type of the button.
          */
diff --git a/core/src/components/segment-button/segment-button.tsx b/core/src/components/segment-button/segment-button.tsx
index a9a5abbd2b..710af58c3f 100644
--- a/core/src/components/segment-button/segment-button.tsx
+++ b/core/src/components/segment-button/segment-button.tsx
@@ -1,10 +1,11 @@
 import type { ComponentInterface } from '@stencil/core';
-import { Component, Element, Host, Prop, State, forceUpdate, h } from '@stencil/core';
+import { Component, Element, Host, Prop, Method, State, forceUpdate, h } from '@stencil/core';
 
 import { getIonMode } from '../../global/ionic-global';
 import type { SegmentButtonLayout } from '../../interface';
 import type { ButtonInterface } from '../../utils/element-interface';
-import { addEventListener, removeEventListener } from '../../utils/helpers';
+import type { Attributes } from '../../utils/helpers';
+import { addEventListener, removeEventListener, inheritAttributes } from '../../utils/helpers';
 import { hostContext } from '../../utils/theme';
 
 let ids = 0;
@@ -26,6 +27,8 @@ let ids = 0;
 })
 export class SegmentButton implements ComponentInterface, ButtonInterface {
   private segmentEl: HTMLIonSegmentElement | null = null;
+  private nativeEl: HTMLButtonElement | undefined;
+  private inheritedAttributes: Attributes = {};
 
   @Element() el!: HTMLElement;
 
@@ -69,6 +72,12 @@ export class SegmentButton implements ComponentInterface, ButtonInterface {
     }
   }
 
+  componentWillLoad() {
+    this.inheritedAttributes = {
+      ...inheritAttributes(this.el, ['aria-label']),
+    };
+  }
+
   private get hasLabel() {
     return !!this.el.querySelector('ion-label');
   }
@@ -87,20 +96,26 @@ export class SegmentButton implements ComponentInterface, ButtonInterface {
     }
   };
 
-  private get tabIndex() {
-    return this.checked && !this.disabled ? 0 : -1;
+  /**
+   * @internal
+   * Focuses the native <button> element
+   * inside of ion-segment-button.
+   */
+  @Method()
+  async setFocus() {
+    const { nativeEl } = this;
+
+    if (nativeEl !== undefined) {
+      nativeEl.focus();
+    }
   }
 
   render() {
-    const { checked, type, disabled, hasIcon, hasLabel, layout, segmentEl, tabIndex } = this;
+    const { checked, type, disabled, hasIcon, hasLabel, layout, segmentEl } = this;
     const mode = getIonMode(this);
     const hasSegmentColor = () => segmentEl?.color !== undefined;
     return (
       <Host
-        role="tab"
-        aria-selected={checked ? 'true' : 'false'}
-        aria-disabled={disabled ? 'true' : null}
-        tabIndex={tabIndex}
         class={{
           [mode]: true,
           'in-toolbar': hostContext('ion-toolbar', this.el),
@@ -119,7 +134,16 @@ export class SegmentButton implements ComponentInterface, ButtonInterface {
           'ion-focusable': true,
         }}
       >
-        <button type={type} tabIndex={-1} class="button-native" part="native" disabled={disabled}>
+        <button
+          aria-selected={checked ? 'true' : 'false'}
+          role="tab"
+          ref={(el) => (this.nativeEl = el)}
+          type={type}
+          class="button-native"
+          part="native"
+          disabled={disabled}
+          {...this.inheritedAttributes}
+        >
           <span class="button-inner">
             <slot></slot>
           </span>
diff --git a/core/src/components/segment/segment.tsx b/core/src/components/segment/segment.tsx
index 4a83c2fa33..f82810f188 100644
--- a/core/src/components/segment/segment.tsx
+++ b/core/src/components/segment/segment.tsx
@@ -165,7 +165,6 @@ export class Segment implements ComponentInterface {
 
   async componentDidLoad() {
     this.setCheckedClasses();
-    this.ensureFocusable();
 
     this.gesture = (await import('../../utils/gesture')).createGesture({
       el: this.el,
@@ -519,19 +518,7 @@ export class Segment implements ComponentInterface {
       const previous = this.checked || current;
       this.checkButton(previous, current);
     }
-    current.focus();
-  }
-
-  /* By default, focus is delegated to the selected `ion-segment-button`.
-   * If there is no selected button, focus will instead pass to the first child button.
-   **/
-  private ensureFocusable() {
-    if (this.value !== undefined) {
-      return;
-    }
-
-    const buttons = this.getButtons();
-    buttons[0]?.setAttribute('tabindex', '0');
+    current.setFocus();
   }
 
   render() {
diff --git a/core/src/components/segment/test/a11y/index.html b/core/src/components/segment/test/a11y/index.html
index 6911c4b1ba..74e4887286 100644
--- a/core/src/components/segment/test/a11y/index.html
+++ b/core/src/components/segment/test/a11y/index.html
@@ -26,6 +26,20 @@
             <ion-label>Shared Links</ion-label>
           </ion-segment-button>
         </ion-segment>
+
+        <br /><br />
+
+        <ion-segment aria-label="Tab Options" color="dark" select-on-focus>
+          <ion-segment-button aria-label="Bookmarks" value="bookmarks">
+            <ion-icon aria-hidden="true" name="bookmark"></ion-icon>
+          </ion-segment-button>
+          <ion-segment-button aria-label="Reading List" value="reading-list">
+            <ion-icon aria-hidden="true" name="glasses"></ion-icon>
+          </ion-segment-button>
+          <ion-segment-button aria-label="Shared Links" value="shared-links">
+            <ion-icon aria-hidden="true" name="link"></ion-icon>
+          </ion-segment-button>
+        </ion-segment>
       </ion-content>
     </ion-app>
   </body>
diff --git a/core/src/components/segment/test/a11y/screen-readers.md b/core/src/components/segment/test/a11y/screen-readers.md
index dfe3a221e3..c36c58215e 100644
--- a/core/src/components/segment/test/a11y/screen-readers.md
+++ b/core/src/components/segment/test/a11y/screen-readers.md
@@ -1,12 +1,12 @@
-"native" refers to this sample: https://w3c.github.io/aria-practices/examples/tabs/tabs-2/tabs.html
+"native" refers to this sample: https://www.w3.org/WAI/ARIA/apg/example-index/tabs/tabs-manual
 
 ### Tabbing to Segment Button
 
 |                          | native                                           | Ionic                                            |
 | ------------------------ | ------------------------------------------------ | ------------------------------------------------ |
-| VoiceOver macOS - Chrome | BOOKMARKS, tab, 1 of 3, Tab Options, tab group   | BOOKMARKS, tab, 1 of 3, Tab Options, tab group   |
+| VoiceOver macOS - Chrome | BOOKMARKS, tab, 1 of 3, Tab Options, tab group   | BOOKMARKS, tab, 1 of 1, Tab Options, tab group   |
 | VoiceOver macOS - Safari | BOOKMARKS, tab, 1 of 3, Tab Options, tab group   | BOOKMARKS, tab, 1 of 3, Tab Options, tab group   |
-| VoiceOver iOS            | Bookmarks, tab                                   | Bookmarks, tab                                   |
+| VoiceOver iOS            | Bookmarks, tab, 1 of 3                           | Bookmarks, tab, 1 of 3                           |
 | Android TalkBack         | Bookmarks, tab                                   | Bookmarks, tab                                   |
 | Windows NVDA             | Tab Options, tab control, BOOKMARKS, tab, 1 of 3 | Tab Options, tab control, BOOKMARKS, tab, 1 of 3 |
 
@@ -14,11 +14,12 @@
 
 |                          | native                                                   | Ionic                    |
 | ------------------------ | -------------------------------------------------------- | ------------------------ |
-| VoiceOver macOS - Chrome | BOOKMARKS, selected, tab, 1 of 3, Tab Options, tab group | BOOKMARKS, selected, tab, 1 of 3, Tab Options, tab group |
+| VoiceOver macOS - Chrome | BOOKMARKS, selected, tab, 1 of 3, Tab Options, tab group | BOOKMARKS, selected, tab, 1 of 1, Tab Options, tab group |
 | VoiceOver macOS - Safari | BOOKMARKS, selected, tab, 1 of 3, Tab Options, tab group | BOOKMARKS, selected, tab, 1 of 3, Tab Options, tab group |
-| VoiceOver iOS            | selected, Bookmarks, tab                                 | selected, Bookmarks, tab |
+| VoiceOver iOS            | selected, Bookmarks, tab, 1 of 3                         | selected, Bookmarks, tab, 1 of 3 |
 | Android TalkBack         | selected                                                 | selected                       |
 | Windows NVDA             | BOOKMARKS, tab, 1 of 3, selected                         | BOOKMARKS, tab, 1 of 3, selected                       |
 
 Note: The `aria-label` for tablist is typically only read on the first interaction.
 
+VoiceOver + Chrome announces the wrong tab count due to the `role="tab"` being in the Shadow DOM: https://bugs.chromium.org/p/chromium/issues/detail?id=1405462
diff --git a/core/src/components/segment/test/a11y/segment.e2e.ts b/core/src/components/segment/test/a11y/segment.e2e.ts
index 085fd51490..f5db2d1614 100644
--- a/core/src/components/segment/test/a11y/segment.e2e.ts
+++ b/core/src/components/segment/test/a11y/segment.e2e.ts
@@ -6,8 +6,7 @@ test.describe('segment: a11y', () => {
   test('should not have any axe violations', async ({ page }) => {
     await page.goto('/src/components/segment/test/a11y');
 
-    // TODO(FW-403): Re-enable rule once segment button is updated to avoid nested-interactive
-    const results = await new AxeBuilder({ page }).disableRules('nested-interactive').analyze();
+    const results = await new AxeBuilder({ page }).analyze();
     expect(results.violations).toEqual([]);
   });