KNOX and BELL transcription factors regulate distinct steps of diploid development in plants. In the green alga Chlamydomonas reinhardtii, KNOX and BELL proteins are inherited by gametes of the opposite mating types and heterodimerize in zygotes to activate diploid development. By contrast, in land plants such as Physcomitrium patens and Arabidopsis thaliana, KNOX and BELL proteins function in meristem maintenance and organogenesis during the later stages of diploid development. However, whether the contrasting functions of KNOX and BELL were acquired independently in algae and land plants is currently unknown. Here, we show that in the basal land plant species Marchantia polymorpha, gamete-expressed KNOX and BELL are required to initiate zygotic development by promoting nuclear fusion in a manner strikingly similar to that in C. reinhardtii. Our results indicate that zygote activation is the ancestral role of KNOX/BELL transcription factors, which shifted toward meristem maintenance as land plants evolved.

20 21 KNOX and BELL transcription factors regulate distinct steps of diploid development in 22 the green lineages. In the green alga Chlamydomonas reinhardtii, KNOX and BELL 23 proteins are inherited by gametes of the opposite mating types, and heterodimerize in 24 zygotes to activate diploid development. By contrast, in land plants such as 25 Physcomitrella and Arabidopsis, KNOX and BELL proteins function in meristem 26 maintenance and organogenesis during the later stages of diploid development. However, 27 whether the contrasting functions of KNOX and BELL were acquired independently in 28 algae and land plants is currently unknown. Here we show that in the basal land plant 29 species Marchantia polymorpha, gamete-expressed KNOX and BELL are required to 30 initiate zygotic development by promoting nuclear fusion in a manner strikingly similar 31 to that of C. reinhardtii. Our results indicate that zygote activation is the ancestral role of 32 KNOX/BELL transcription factors, which shifted toward meristem maintenance as land 33 plants evolved. 34 35 106 6 the functional transition of KNOX/BELL from zygote activation to sporophyte 107 morphogenesis occurred at least once in the land plant lineage independently of the 108 acquisition of multicellular sporophytes. Additionally, we uncovered inverted sex-109 specific expression patterns of KNOX and BELL genes between C. reinhardtii and M. 110 polymorpha, suggesting that anisogamy evolved independently of KNOX/BELL 111 expression in gametes.112 113 Results 114MpKNOX1 is as an egg-specific gene in M. polymorpha 115 We previously reported that an RWP-RK TF MpRKD promotes egg cell differentiation 116 in M. polymorpha. Loss-of-function Mprkd mutant females grow normally and produce 117 archegonia like the wild type, but their egg cells do not mature, instead degenerating after 118 ectopic cell division and vacuolization (Koi et al., 2016). We made use of this egg-specific 119 defect in Mprkd to identify genes preferentially expressed in egg cells of M. polymorpha. 120 Briefly, we collected ~2,000 archegonia from two independent Mprkd female mutant 121 lines (Mprkd-1 and Mprkd-3, Koi et al., 2016), each in two replicates. As a control, 122 ~4,000 archegonia were collected from wild-type females in four replicates. We extracted 123 RNA from each pool and analyzed it by next-generation sequencing. Comparative 124 transcriptome analysis identified 1,583 genes with significantly reduced mRNA levels in 125 Mprkd compared to wild-type archegonia (>3-fold and false discovery rate < 0.01; Figure 126 1A and Figure1 -figure supplement 1A). Among these, MpKNOX1 (Mp5g01600), the 127 only class I KNOX gene in M. polymorpha (Bowman et al., 2017, Frangedakis et al., 128 2017), was strongly downregulated in Mprkd vs. the wild type (~500-fold) (Figure 1B). 129 7 The MpKNOX1 polypeptide contains KNOX I, KNOX II, ELK, and Homeobox domains, 130as do KNOX proteins from green algae, mosses, ferns and flowering plants ( Figure 1C). 131Previous RNA-sequencing data (Bowman et al., 2017) indi...

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The Bombyx mori latent virus (BmLV) belongs to the unassigned plant virus family Tymoviridae and contains a positive-sense, single-stranded RNA genome. BmLV has infected almost all B. mori-derived cultured cell lines through unknown routes. The source of BmLV infection and the BmLV life cycle are still unknown. Here, we examined the interaction between BmLV and the insect DNA virus Bombyx mori nucleopolyhedrovirus (BmNPV). Persistent infection with BmLV caused a slight delay in BmNPV propagation, and BmLV propagation was enhanced in B. mori larvae via co-infection with BmNPV. We also showed that BmLV infectious virions were co-occluded with BmNPV virions into BmNPV occlusion bodies. We propose a new relationship between BmLV and BmNPV.