Exploring potential strategies for haploid induction based on double fertilization in plants

Abstract

Haploid induction (HI), an indispensable procedure in doubled haploid breeding, has attracted increasing attention in crop genetic improvement due to its ability to rapidly fix desirable traits in a homozygous state, thereby shortening the breeding cycle. However, HI has only been successfully implemented in a limited number of crops, and its underlying mechanisms remain largely enigmatic. This review summarizes five potential HI routes based on previous findings and the key events during the process of double fertilization in flowering plants. Among these HI methods, we suggest that sperm DNA fragmentation and ectopic expression of embryogenesis activator, as straightforward avenues for discovering new HI-related genes. We also emphasize that the combination of genome editing techniques with HI is a promising strategy to accelerate crop improvement and doubled haploid breeding. We envision that the proposed directions can pave the way for improving and deepening our understanding of HI mechanisms.

Keywords: double fertilization; ectopic expression; haploid induction; membrane fusion; sperm DNA fragmentation; uniparental chromosome elimination.

Figure 1

Hypothetical mechanistic model of HI according to double fertilization in plants. (a) Diagram of an unfertilized ovule showing its eight constituent nuclei. (b) During the first stage of double fertilization, after the pollen tube ruptures, sperm cells are released and enter the embryo sac. (c) First method for HI (H1): Elevated reactive oxygen species (ROS) levels result in the fragmentation of sperm DNA and trigger HI. (d) Communication and fusion between the plasma membranes of the male and female gametes. (e) Second method for HI (H2): A haploid state is induced by disrupting plasma membrane communication (requiring KOKOPELLI [KPL]) or fusion (requiring DOMAIN OF UNKNOWN FUNCTION 679 PROTEIN [DMP]). (f) Fusion of the nuclear membranes from the sperm cell and the egg cell. (g) Third method for HI (H3): Blocking the fusion of the nuclear membranes results in the degradation of the sperm nucleus. (h) Chromosome movement and separation before initiation of zygote development. (i) Fourth method for HI (H4): CENTROMERIC HISTONE H3 (CENH3) function is directly (with cenh3 mutants) or indirectly (with mutation of KINETOCHORE NULL 2 [KNL2]) disturbed, causing the elimination of chromosomes from the maternal or paternal genome during early embryogenesis. (j) Activation of embryogenesis. (k) Fifth method for HI (H5): Ectopic expression of BABY BOOM (BBM), PARTHENOGENESIS (PAR), or WUSCHEL (WUS) leads to advanced activation of egg cells to generate haploids with only the maternal genome.

Figure 2

Haploid induction via sperm DNA fragmentation and elevated ROS levels. The mutation of genes related to ROS levels (for example, MTL/NLD/PLA1, POD65, or PLD3) or treatment of pollen grains with ROS‐inducing reagents (like phosphatidylcholine [PC] and methimazole [MMI]) induces DNA fragmentation in sperm cells and triggers haploid induction.

Figure 3

Haploid induction via disrupted communication and fusion between the plasma membranes of the male and female gametes.

Figure 4

Haploid induction via blocking the fusion of the male and female nuclear membranes. Total degradation of paternal chromosomes will result in haploidy, while partial chromosome degradation will lead to the appearance of chimeric embryos.

Figure 5

Haploid induction via the CENH3‐HI system leading to uniparental chromosome elimination. CENH3 function is directly (cenh3) or indirectly (knl2) disturbed, causing the elimination of maternal or paternal chromosomes during early embryogenesis.

Figure 6

Haploid induction via the ectopic expression of BBM, PAR, or WUS, leading to advanced independent activation of egg cells to generate haploids without the paternal genome.