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

13 14 • Population divergence in early development may lead to hybrid developmental failure and 15 inviability, and may be a factor driving speciation in flowering plants. 16 • We investigate the pattern of early and late embryo and endosperm development of seed 17 from Mimulus guttatus and the closely related, serpentine endemic M. nudatus, and compare 18 it to that of hybrid seed derived from reciprocal, interspecific crosses. We address whether 19 disruption in hybrid seed development is the primary source of reproductive isolation 20 between these sympatric taxa. 21 • M. guttatus and M. nudatus differ in the pattern and timing of endosperm and embryo 22 development. Some hybrid seed exhibit early disruption of endosperm development and are 23 completely inviable, while other hybrid seed exhibit comparatively normal early 24 development, but show impaired endosperm proliferation. These developmental patterns 25 underlie the phenotypes of mature seed, which are either very small and flat, (indicating little 26 to no endosperm), or shriveled, (indicating reduced endosperm volume). Both phenotypes 27 have low germination success. Hybrid seed inviability forms a potent reproductive barrier 28 between M. guttatus and M. nudatus. 29 • We provide a partial developmental mechanism for the hybrid barrier between M. guttatus 30 and M. nudatus. In addition to illustrating the early stages of endosperm and embryo 31 development, we shed light on the extent of developmental variation between closely related 32 species within this important ecological model system. 33 34 39 Introduction 40 41 The process of gradual evolution imposes a fundamental constraint on organismal 42 development – each successful evolutionary shift, large or small, must allow for viable offspring 43 44 by the disruption of development frequently observed when two divergent populations hybridize, 45 when both lineages themselves continue to produce viable offspring. In nature, these 46 incompatibilities can keep species distinct by preventing gene flow. In the laboratory, we can 47 make use of incompatibilities witnessed in hybrid offspring to investigate how development has 48 evolved in isolation and how evolutionary constraint may shape developmental trajectories. Here 49 we describe differences in seed development between a recently diverged species pair – Mimulus 50 guttatus and M. nudatus. We further show that postzygotic failures of development are largely 51 responsible for incompatibility in experimental crosses between this sympatric species pair. We 52 propose that the M. guttatus sp. complex may serve as a new model to …