ARTICLE Multispecies autocatalytic RNA reaction networks in coacervates Sandeep Ameta 1,3 ✉ , Manoj Kumar 1 , Nayan Chakraborty 1 , Yoshiya J. Matsubara 1 , Prashanth S 1 , Dhanush Gandavadi 1 & Shashi Thutupalli 1,2 ✉ Robust localization of self-reproducing autocatalytic chemistries is a key step in the realization of heritable and evolvable chemical systems. While autocatalytic chemical reaction networks already possess attributes such as heritable self-reproduction and evolvability, localizing functional multispecies networks within complex primitive phases, such as coacervates, has remained unexplored. Here, we show the self-reproduction of the Azoarcus ribozyme system within charge-rich coacervates where catalytic ribozymes are produced by the autocatalytic assembly of constituent smaller RNA fragments. We systematically demonstrate the catalytic assembly of active ribozymes within phase-separated coacervates—both in micron-sized droplets as well as in a coalesced macrophase, underscoring the facility of the complex, charge- rich phase to support these reactions in multiple configurations. By constructing multispecies reaction networks, we show that these newly assembled molecules are active, participating both in self- and cross-catalysis within the coacervates. Finally, due to differential molecular transport, these phase-separated compartments endow robustness to the composition of the collectively autocatalytic networks against external perturbations. Altogether, our results establish the formation of multispecies self-reproducing reaction networks in phase-separated compartments which in turn render transient robustness to the network composition. https://doi.org/10.1038/s42004-023-00887-5 OPEN 1 Simons Centre for the Study of Living Machines, National Centre for Biological Sciences, Tata Institute of Fundamental Research, Bengaluru, Karnataka, India. 2 International Centre for Theoretical Sciences, Tata Institute of Fundamental Research, Bengaluru, Karnataka, India. 3 Present address: Trivedi School of Biosciences, Ashoka University, Plot No. 2, Rajiv Gandhi Education City, P.O. Rai, Sonepat, Haryana 131029, India. ✉ email: sandeep.ameta@ashoka.edu.in; shashi@ncbs.res.in COMMUNICATIONS CHEMISTRY | (2023)6:91 | https://doi.org/10.1038/s42004-023-00887-5 | www.nature.com/commschem 1 1234567890():,;