<p>by Rebecca E. Dale, Silke Berger, Sara Alaei, Marcus C. Davis, Frank J. Tulenko, Peter D. Currie</p> The origin of paired fins was a major event in early vertebrate history that fueled the adaptive radiation of the gnathostome clade. Evidence for the conservation of ancient regulatory systems between paired and median fins supports a model in which appendage-patterning mechanisms first evolved in the midline and were later redeployed to a new embryonic context, the flank. However, while paired fins and limbs are asymmetric along the dorsoventral (DV) axis, the equivalent axis does not exist in median fins, which are symmetric across the midline, raising questions of how DV patterning evolved. Here, we combine genetic tools in zebrafish with comparative expression analyses in representative ray-finned and cartilaginous fishes to show that a subset of limb DV patterning genes are expressed during median fin development. Using genetic lineage tracing of the canonical limb dorsalizing factor <i>lmx1bb</i>, we further demonstrate that, although <i>lmx1bb</i>-derived cells occupy distinct spatial domains across fin types, a subset of this lineage is fated to differentiate into fin ray osteoblasts in both paired and median fins. To test the potential conservation of regulatory modules across fin types, we generated a series of deletion mutants for two known <i>lmx1b</i> mouse limb enhancers, <i>LARM1</i> and <i>LARM2,</i> that we show are partially conserved upstream of zebrafish <i>lmx1bb</i>. These experiments revealed that <i>LARM</i> function is essential for dorsoventral patterning of the paired pectoral and pelvic fins of zebrafish but is dispensable for formation of the unpaired dorsal and anal fins. Collectively, these data support the conservation of an ancient <i>LARM</i>-Lmx1b dorsal determination system in the paired fins of osteichthyans, challenge models proposing simple co-option of a pre-existing median fin gene regulatory network and suggest that the acquisition of this regulatory module may have been a key early step in the evolution of the paired fin DV patterning axis.