The Himalaya–Hengduan Mountain (HHM) region consists of two global biodiversity hotspots characterized by a high degree of plant endemism. However, little is known about how these endemic species are formed and maintained in relation to the regional geomorphology of the past or current time. Thus, this study investigated the genetic structure of the herbaceous genus Acanthocalyx (Caprifoliaceae) endemic to the HHM to demonstrate if major geographic or ecological barriers in the HHM region have influenced its phylogeographic patterns. Our analyses revealed distinct genetic structures within A. alba and A. nepalensis and indicated that A. delavayi may have recently evolved from isolated peripheral populations of A. nepalensis. In particular, we not only confirmed a well-known genetic structure of alpine plants between the Himalayas and the Hengduan Mountains but also discovered a notable floristic boundary (bounded by 30° to 31°N latitude) within the Hengduan Mountains from A. alba. This study provides new insights into the dispersal and intraspecific genetic variation of Acanthocalyx and highlights the importance of geomorphological features for the diversification of HHM alpine flora.