Poster Presentation The Annual Scientific Meeting of the Endocrine Society of Australia and the Society for Reproductive Biology 2014

The lncRNA HOTTIP during phallus development in a marsupial (#317)

Hongshi Yu 1 , Keng Yih Chew 1 2 , Marilyn B Renfree 1
  1. Department of Zoology, The University of Melbourne, Melbourne, Victoria 3010, Australia
  2. Institute of Biology, University of Leiden, Leiden, the Netherlands

HOTTIP (HOXA transcript at the distal tip) maintains active chromatin to coordinate homeotic gene expression. Depleted HOTTIP leads to shortening of distal bony elements in chicken forelimbs. HOTTIP is involved in cartilage integrity and also associated with hepatocellular carcinoma. However, most of its role in controlling cell differentiation, development and growth remains largely unknown. The molecular network of key genes controlling development of the appendages, the limbs and phallus, are very similar. Since HOTTIP regulates HOX gene expression especially in the HOXA13, it is reasonable to expect that HOTTIP may participate in appendage development. We therefore investigated the evolution of the long non-coding RNA (lncRNA) HOTTIP and its role in the network controlling external genitalia development in a marsupial, tammar wallaby.

LncRNAs are rapidly evolving genes and are poorly conserved in most cases. We therefore employed the Infernal program (INFERence of RNA ALignment) and chromosome localization to identify partially conserved regions and sequenced both 5’- and 3’- ends produced by RACE PCR to obtain the full-length of tammar HOTTIP. We sampled limb and phallus tissues from full term fetuses, and from pouch young aged up to 50 days post-partum, covering the critical time when the phallus becomes sexually dimorphic in the tammar. As expected, HOTTIP was expressed in both developing limbs and phallus. To further investigate its potential role, whole mount in situ hybridization (WISH) was performed and showed that tammar HOTTIP and HOXA13 were co-localised in both limb and phallus, inferring that tammar HOTTIP may regulate HOXA13 expression as in the human. This is the first demonstration that the long non-coding RNA HOTTIP may be involved in external genitalia development in a marsupial.

HY and KYC contributed equally to this study