Title : Disruption of endosperm development is a major cause of hybrid seed inviability between 1

13 14 • Divergence of developmental mechanisms within populations may lead to hybrid 15 developmental failure, and may be a factor driving speciation in angiosperms. 16 • We investigate patterns of endosperm and embryo development in Mimulus guttatus and the 17 closely related, serpentine endemic M. nudatus, and compare them to those of reciprocal 18 hybrid seed. We address whether disruption in hybrid seed development is the primary 19 source of reproductive isolation between these sympatric taxa. 20 • M. guttatus and M. nudatus differ in the pattern and timing of endosperm and embryo 21 development. Some hybrid seed exhibit early disruption of endosperm development and are 22 completely inviable, while others develop relatively normally at first, but later exhibit 23 impaired endosperm proliferation and low germination success. These developmental 24 patterns are reflected in mature hybrid seed, which are either small and flat (indicating little 25 to no endosperm), or shriveled (indicating reduced endosperm volume). Hybrid seed 26 inviability forms a potent reproductive barrier between M. guttatus and M. nudatus. 27 • We shed light on the extent of developmental variation between closely related species 28 within the M. guttatus species complex, an important ecological model system, and provide a 29 partial mechanism for the hybrid barrier between M. guttatus and M. nudatus. 30 31 Introduction 42 The process of gradual evolution imposes a fundamental constraint on organismal 43 development – each successful evolutionary shift, large or small, must allow for viable offspring 44 45 by the disruption of development frequently observed when two divergent populations hybridize, 46 when both lineages themselves continue to produce viable offspring. In nature, these 47 incompatibilities can keep species distinct by preventing gene flow. In the laboratory, we can 48 make use of incompatibilities witnessed in hybrid offspring to investigate how development has 49 evolved in isolation and how evolutionary constraint may shape developmental trajectories. Here 50 we describe differences in seed development between a recently diverged species pair – Mimulus 51 guttatus and M. nudatus. We further show that postzygotic failures of development are largely 52 responsible for incompatibility in experimental crosses between this sympatric species pair. We 53 propose that the M. guttatus sp. complex may serve as a new model to understand the evolution 54 of development and developmental abnormalities in hybrid plants. 55 56 Development in multicellular organisms requires coordination across numerous cell lineages or 57 types. The process …