Plants come from seeds. This happens when the seeds are planted in the ground and sprout (begin to grow). Before a seed can sprout, it must go through a process called germination. The process of germination happens inside the seed. To learn more about the process of germination, let’s take a look inside a seed…
The parts of a seed
Before we look at the inside of a seed, let’s talk about the outside of the seed. The outside of a seed is called the seed coat. The seed coat is the hard outer layer of the seed. It is the part we see and hold in our hands before we plant them in the ground or a pot of soil.
Not all seed coats are alike, though. Some are hard (corn, beans, peas, okra, morning glories). Other seeds have soft seed coats (marigolds, tomatoes, zinnias, peppers, cucumbers).
The inside of a seed has four main parts. The four main parts of the inside of a seed are:
- The Epicotyl
- The Hypocotyl
- The Radicle
- The Cotyledon
Now let’s look at what each of these parts becomes once the seed becomes a plant.
The Epicotyl are the parts of the seed that become the first leaves of a plant.
The Hypocotyl is the stem of the plant.
The Radicle is the first root the plant has.
The Cotyledon is the inner protective layer of the seed that stores food for the seed to use during the process of germination and until the seed comes through the soil and has leaves that can be used for photosynthesis.
The process of germination
If you have ever planted a seed, you know how exciting it is to see the plant that comes from that seed break through the soil. Have you ever thought about how it happens? Let’s find out!
When you plant seeds in some soil, it is important to keep the soil watered (not too much). The reason this is so important is because the seeds you plant need to be able to take in oxygen and minerals from the soil and water through the seed coat’s tiny pores (holes) to give the inside of the seed the food it needs to break open and make its way through the soil so it can grow into a plant.
When the seed is full enough, it pops open. The first parts of the seed to come through the seed coat are the cotyledon and the radicle (root). The root takes hold of the soil and starts to take in food from the soil. But because it is still so small, the cotyledon is still the main source of food for the seed.
The next part of the seed that appears is the hypocotyl. The hypocotyl is sometimes called the understem because it first appears under the cotyledon. The hypocotyl continues to grow upward with the epicotyl. The epicotyl becomes the first leaves of the new plant.
By the time the epicotyl are showing, the plant is now above the ground. When this happens, the cotyledon (which is sometimes called the seed leaves and looks like thin, dried brownish-white skin) has finished its job. Because their job is done, they fall off the plant and become part of the soil.
Once the cotyledon are gone, the plant’s tiny leaves take over the job of supplying food to the new plant. And that is the process called germination.
All seeds are not alike
If you look at different kinds of seeds, you can easily see that they are not all alike. Seeds come in different sizes, shapes and colors And like you’ve already learned, some seeds have softer seed coats than others. All these differences mean that seeds germinate differently.
Seeds with hard seed coats usually germinate slower than seeds with soft seed coats. Why do you think this is?
The reason seeds with hard seed coats take longer to germinate is that it takes longer for the seed to drink enough water to soften the seed coat enough that the inside parts of the seed can break through.
There are also other reasons some seeds take longer to germinate than others. Here are a few of them:
- The amount of sunshine. Seeds don’t see the sun, but the sun heats the soil to make it warm and cozy—which is just what a seed needs to germinate.
- The amount of water in the ground. If the soil is too dry, the seed cannot get the water it needs. If it is too wet, the ground will not have enough oxygen in it to give the seed what it needs to germinate.
- Planting the seed too deep. If you plant a seed too deep, it will use all the energy and food stored in the cotyledon before it can break through the ground so the leaves can come out and take over feeding the plant.
- The seasons. Most seeds will not germinate in the fall or winter. The ground is too cold during these two seasons for a seed to germinate. Instead, the seeds sleep until spring. When a seed sleeps, it is dormant.
Planting seeds and watching them grow is one of the most exciting things ever!
Fill each section of an empty egg carton with moist potting soil. Place a different kind of seed in each section. Make a chart showing what seeds are planted in each section of the egg carton. Keep the seeds in a warm, sunny place and keep the soil moist—but not too wet. Write down how many days it takes for each seed to germinate and pop through the soil.
Genes and or factors that induce seed dormancy or inhibit germination
The first important stage for dormancy induction is probably the end of the morphogenetic program, when all tissues present in a mature embryo have been formed and the embryo enters a phase of growth arrest. ABA-INSENSITIVE3 (ABI3), FUSCA3 (FUS3) and LEAFY COTYLEDON (LEC1 and LEC2) are four key regulators that play prominent roles in controlling mid- and late seed development (Meinke et al., 1994; Raz et al., 2001). ABI3, FUS3 and LEC2 encode related plant-specific transcription factors containing the conserved B3 DNA binding domain (Giraudat et al., 1992; Luerssen et al., 1998; Stone et al., 2001), whereas LEC1 encodes a HAP3 subunit of the CCAAT binding transcription factor (CBF, also known as NF-Y (Lotan et al., 1998)). All four abi3, lec1, lec2 and fus3 mutants are severely affected in seed maturation and share some common phenotypes, such as decreased dormancy at maturation (Raz et al., 2001) and reduced expression of seed storage proteins (Gutierrez et al., 2007). However, they also show specific phenotypes, such as the absence of chlorophyll degradation in the dry seed (abi3), a reduced sensitivity to ABA (abi3 and, to a lesser extent, lec1), the accumulation of anthocyanins (fus3, lec1, and, to a lesser extent, lec2), an intolerance to desiccation (abi3, fus3, and lec1), or defects in cotyledon identity (lec1, fus3, and lec2) (Bäumlein et al., 1994; Keith et al., 1994; Meinke et al., 1994; Parcy et al., 1994; Parcy & Giraudat, 1997; Luerssen et al., 1998; Vicient et al., 2000; Raz et al., 2001; Stone et al., 2001; Kroj et al., 2003). It was recently shown that several of the fus3 phenotypes are due to pleiotropic effects caused by truncated gene products of the mutant alleles. The direct effects of FUS3 are probably restricted to embryo-derived dormancy and determination of cotyledon epidermis cell identity (Tiedemann et al., 2008).
The LEC1 gene is required for normal development during early and late phases of embryogenesis and is sufficient to induce embryonic development in vegetative cells (Lotan et al., 1998). Loss of function of LEC1 leads to germination of excised embryos at a similar stage (between 8–10 days after pollination) as lec2 and fus3 mutants, but earlier during embryo development than found for abi3 mutants (Raz et al., 2001). Ten HAP3 (AHAP3) subunits have been identified in Arabidopsis, which can be divided into two classes based on sequence identity in their central, conserved B domain (Kwong et al., 2003). LEC1 and the closely related subunit, LEC1-LIKE (L1L), constitute LEC1-type AHAP3 subunits, whereas the remaining eight are designated non-LEC1-type. Similar to LEC1, L1L is expressed primarily during seed development. However, suppression of L1L gene expression induced defects in embryo development that differed from those of lec1 mutants, suggesting that LEC1 and L1L play different roles in embryogenesis (Kwong et al., 2003).
LEC2 directly controls a transcriptional program involved in the maturation phase of seed development. Induction of LEC2 activity in seedlings causes rapid accumulation of RNAs normally present primarily during the maturation phase, including seed storage and lipid-body proteins. Promoters of genes encoding these maturation RNAs all possess RY motifs (cis-elements bound by B3 domain transcription factors) (Braybrook et al., 2006). This provides strong evidence that these genes represent transcriptional targets of LEC2. One of these genes is DOG1, the first seed dormancy gene accounting for variation occurring in natural populations that has been identified at the molecular level (Bentsink et al., 2006).
It has been shown that ABI3, FUS3, LEC1 and LEC2 interact as a network to control various aspects of seed maturation. LEC1 was shown to regulate the expression of both ABI3 and FUS3 (Kagaya et al., 2005), FUS3 and LEC2 have been shown to act in a partially redundant manner to control gene expression of seed specific proteins, and LEC2 was shown to locally regulate FUS3 expression in regions of the cotyledons (Kroj et al., 2003). The indication of redundant regulation within this group of genes was recently shown (To et al., 2006). By analyzing ABI3 and FUS3 expression in various single, double, and triple maturation mutants, multiple regulatory links among all four genes were identified. It was found that one of the major roles of LEC2 was to up-regulate FUS3 and ABI3. The lec2 mutation leads to a dramatic decrease in ABI3 and FUS3 expression, and most lec2 phenotypes can be rescued by ABI3 or FUS3 constitutive expression. In addition, ABI3 and FUS3 were shown to positively regulate themselves and each other, thereby forming feedback loops essential for their sustained and uniform expression in the embryo. Finally, LEC1 also positively regulates ABI3 and FUS3 in the cotyledons (To et al., 2006). Although multiple regulatory links were identified amongst these four genes, molecular mechanisms underlying this network, and the downstream targets of the network associated with dormancy induction still require further investigation.
Apart from mutants that influence general seed maturation, other mutants more specifically influence seed dormancy, i.e. mutants, which are altered in ABA biosynthesis or its mode of action. ABA regulates various aspects of plant growth and development, including seed dormancy. The absence of ABA-induced dormancy allows seeds to germinate without gibberellins. Therefore, the selection of mutants that germinate in the presence of GA biosynthesis inhibitors, such as paclobutrazol and tetcyclacis, is an effective way to isolate ABA biosynthesis mutants (Léon-Kloosterziel et al., 1996b). Reciprocal crosses between wild type and the ABA deficient aba1 mutants showed that dormancy is controlled by the ABA genotype of the embryo and not by that of the mother plant. The latter is responsible for the relatively high ABA levels found in seeds halfway through seed development (Karssen et al., 1983). At this phase ABA may prevent precocious germination as shown by the maternal ABA effects in the extreme aba abi3-1 double mutants (Koornneef et al., 1989). Key genes for ABA biosynthesis during seed development are NCED6 and NCED9, both members of the 9-cis-epoxycarotenoid dioxygenese family (Lefebvre et al., 2006). In contrast to what was proposed by Karssen et al (1983) it is now clearly shown that endogenous ABA is required for the maintenance of seed dormancy. In wild type ABA levels decrease at the end of seed maturation and during imbibition due to the activity of ABA catabolism genes belonging to the P450 CYP707A family (Okamoto et al., 2006), indicating that ABA levels can be modified at different phases of seed development and germination with significant effects on germination. Furthermore, the observation that inhibitors of ABA biosynthesis, such as nor-fluorazon, promote germination (Debeaujon and Koornneef, 2000) indicated that the maintenance of dormancy in imbibed seeds is an active process involving de novo ABA synthesis as was also shown for dormant seed batches of the accession Cvi (Ali-Rachedi et al., 2004).
ABA has a major effect on seed dormancy and therefore it can be expected that defective ABA signalling also leads to changes in germination characteristics. Substantial progress has been made in the characterization of such ABA signal transduction pathways (Bonetta and McCourt, 1998; Leung and Giraudat, 1998). Genetic screens to identify ABA signalling mutants were based primarily on the inhibition of seed germination by applied ABA. The ABA-insensitive (abi) mutants (Koornneef et al., 1984) and several others described thereafter (Rock, 2000 and Holdsworth et al., 2008a) are able to germinate and grow in the presence of ABA concentrations that are inhibitory to the wild type. It was expected that such screens would yield ABA receptor and signal transduction mutants. However most of ABA receptor genes were identified using reverse genetics in which screening for germination characteristics were performed that often showed no or small effects on ABA sensitivity for germination and dormancy it self was often not tested (reviewed in Holdsworth et al., 2008a). Forward screens for mutants in which seed germination is prevented by low concentrations of ABA that ordinarily permit germination of the wild-type seed were first described by Cutler et al. (1996) resulting in the era1 (enhanced response to ABA) to era3 mutants. Subsequently using similar screens identified many additional loci that are involved in removal of sensitivity to ABA function, that when mutated lead to ABA hypersensitivity of imbibed after-ripened or moist-chilled seeds (Hugouvieux et al., 2001; Xiong et al., 2001; Nishimura et al., 2004; Katagiri et al., 2005; Zhang et al., 2005; Pandey et al., 2006; Saez et al., 2006; Yoine et al., 2006; Nishimura et al., 2007). These loci encode diverse functions, including those associated with RNA translation and metabolism, protein degradation pathways and phosphatase components of signalling pathways and transcription factors (Holdsworth et al., 2008a). As judged from their effects on seed dormancy, these two sets of mutations also alter the regulation of seed germination by endogenous ABA. The abi3, abi4 and abi5 mutants exhibit reduced expression of various seed maturation genes but only abi3 mutants are non-dormant, which coincides with desiccation intolerance (Nambara et al., 1992, Ooms et al., 1993, Bies et al., 1999, Finkelstein et al., 2008) in strong alleles. Surprisingly no dormancy or other seed maturation phenotype was observed in abi4 and abi5 mutants (Finkelstein, 1994; Finkelstein et al., 2008), except reduction of some seed maturation specific mRNAs (Finkelstein and Lynch, 2000; Söderman, et al., 2000). This may indicate that other genes are redundant in function to these seed specific transcription factors, which are members of the APETALA2 domain (ABI4, Finkelstein et al., 1998; Söderman, et al., 2000) and basic leucine zipper factor family (ABI5, Finkelstein and Lynch, 2000; Lopez-Molina et al., 2001).
According to a recent report, ABA levels might be positively regulated by DELLA protein through upregulation of XERICO expression (Zentella et al., 2007). XERICO over-expression, which encodes an E3 ubiquitin ligase, leads to both an elevated level of ABA and increased drought tolerance, although the mechanism is still unknown (Ko et al., 2006).
A class of mutants that was directly selected on the basis of reduced dormancy are the rdo1-rdo4 mutants (Léon-Kloosterziel et al., 1996a; Peeters et al., 2002). The fact that all four mutants show some mild pleiotropic effects in adult plants indicates that the genes are not specific for dormancy/germination but affect other processes as well. The RDO4 (renamed as HUB1) was shown to encode a C3HC4 Ring finger protein involved in the monoubiquitination of histone H2B, revealing a role for chromatin modelling in seed dormancy (Liu et al., 2007). The dag1-1 mutant also displays reduced dormancy, but in contrast to the rdo mutants the effect is determined by the maternal genotype. This is in agreement with the expression pattern of the DAG1 gene in the vascular tissue of the developing seed. DAG1, which encodes a DOF transcription factor, may influence the import of compounds from the mother plant into the seed (Papi et al., 2000). It is the first gene identified as being specifically involved in maternal control of seed germination. However, the germination phenotype of dag2, mutant in the related DAG2 gene, with a similar expression pattern as DAG1 is opposite to that of dag1 seeds (Gualaberti et al., 2002) showing increased dormancy. Additional mutants with a reduced dormancy phenotype at other loci, including mutants with no obvious pleiotropic effect have been isolated (Bentsink et al., 2006 and M. Schwab and W. Soppe unpublished results), indicating the complexity of the genetic regulation of seed dormancy. Instead of selecting for mutants that germinate when the wild type is still dormant, Salaita et al. (2005) used germination speed at 10°C as a selection criterion when screening activation tagged lines of the Col accession. Except for two tt mutants (see below) none of these cold temperature germination (ctg) has been characterized molecularly.
Another group of mutants that shows reduced seed dormancy are mutants with an altered seed coat or testa (Debeaujon et al., 2000; reviewed in Debeaujon et al., 2007; Lepiniec et al., 2006). The seed coat is a multifunctional organ that plays an important role in embryo nutrition during seed development and in protection against detrimental agents from the environment afterwards (Debeaujon et al., 2007). The seed coat is formed from two integuments of epidermal origin that surround the mature ovule. The development of the seed coat from the ovule has been described by Beeckman et al. (2000).
The seed coat together with the endosperm layer exerts a germination-restrictive action, either by being impermeable to water and/or oxygen, by producing germination inhibiting compounds or by its mechanical resistance to radicle protrusion. In Arabidopsis, phenolic compounds and their derivatives present in the inner layer of the testa, called endothelium, affect seed coat properties that influence germination as can be concluded from the reduced dormancy phenotype of many testa mutants.
Seed coat mutants consist of two major groups. One group, affected in flavonoid pigmentation is represented by the transparent testa (tt) and transparent testa glabra (ttg) mutants. Mutants identified are tt1 to tt15, ttg1 and ttg2 and banyuls (ban). The color of the tt mutants ranges from yellow to pale brown (Debeaujon et al., 2000). Ban mutants accumulate pink flavonoid pigments in the endothelium of immature seeds, but do not contain proanthocyanins due to a mutation in the anthocyanin reductase (ANR) gene (Xie et al., 2003) resulting in grayish-green, spotted mature seeds (Albert et al, 1997; Devic et al, 1999). The ttg1 mutant lacks mucilage and trichomes and is affected in the morphology of the outer layer of the seed coat as well as in pigment production. Many of the seed coat mutants have now been cloned, for more details see supplemental Table 1 online and Debeaujon et al. (2007).
The second group is represented by mutants affected in testa structure. The aberrant testa shape (ats) mutant, mutated in the KANADI 4 gene (McAbee et al., 2006) produces a single integument instead of the two integuments seen in wild-type ovules and shows less dormancy.
Genes and or factors that decrease seed dormancy or promote the germination potential of seeds
The germination of Arabidopsis seeds is under phytochrome-mediated photocontrol. It is therefore to be expected that phytochrome deficient mutants are affected in seed germination. The complexity of the phytochrome system comes from the presence of distinct types of phytochromes, for which five genes in the Arabidopsis genome encode different, but related, apoproteins (Sharrock and Quail, 1989). In addition different modes of action of phytochrome, described as very-low-fluence response (VLFR), low-fluence response (LFR) and high-irradiance response (HIR), which have their own fluence dependency, can affect germination (reviewed by Casal and Sánchez, 1998). Mutants lacking phytochrome B (phyB) show a reduced sensitivity to red light, indicating that phyB has a primary role in seed germination. PhyA can only induce germination after a prolonged imbibition of seeds (Shinomura et al., 1994). Detailed action spectra for seed germination performed in wild type, phyA and phyB mutants revealed a typical red/far-red (R/FR) -reversible LFR mediated by phyB, whereas the germination response mediated by phyA turned out to be a VLFR with a 104 -fold higher sensitivity to light (Shinomura et al, 1996). The observation that also phyA phyB double mutants show some light- dependent germination indicates the involvement of another R/FR-reversible photoreceptor system (Yang et al., 1995; Poppe and Schäfer, 1997) probably mediated by phyC, D, and/or E.
Although the main role of phytochrome is in light-induced stimulation of seed germination, a role in the onset of dormancy or the setting of the light requirement is suggested by the experiments of McCullough and Shropshire (1970) and Hayes and Klein (1974). These authors showed that the R/FR ratio experienced by the mother plant and therefore during seed maturation, affects the subsequent germination behaviour of mature seeds. Munir et al. (2001) showed that photoperiod conditions during seed formation may also influence seed germination. However, this effect was strongly genotype dependent. In addition, it appears that phytochrome mediated pathways are required to break cold-induced dormancy (Donohue et al., 2007). Cool temperatures during seed maturation induced seed dormancy which could not be overcome in the hy2-1 (deficient in phytochrome chromophore, common to all five phytochromes) mutant.
The plant hormone gibberellin plays an important role in promoting seed germination. GA-deficient mutants are unable to germinate without exogenous GAs (Koornneef and van der Veen, 1980; Mitchum et al., 2006). De novo biosynthesis of GAs is required during imbibition, as was concluded from the observation that inhibitors of GA biosynthesis, such as paclobutrazol and tetcyclacis prevent germination (Karssen et al., 1989) unless ABA is absent. As expected also GA signalling mutants such as the sleepy1 (sly1) mutant, which was selected in a screen for suppressors of the ABA insensitive mutant abi1-1 (Steber et al., 1998) and Della protein encoding genes such as RGL2 (Lee et al., 2002; Bassel et al., 2004) have germination defective phenotypes. For the GA receptors mutants (gid1a, gid1b, gid1c) triple mutants had to be constructed to see this phenotype due to redundancy of the function of these genes (Griffith et al., 2006, Iuchi et al., 2007, reviewed by Hirano et al., 2008). GAs can promote germination by their ability to overcome germination constraints that exist in seeds requiring after-ripening, light and cold. This led to the suggestion that such environmental factors may induce GA biosynthesis during the early phases of germination. At present the changes of GA content and the expression of GA biosynthesis and catabolism genes during dormancy release and germination is well documented (Yamauchi et al., 2007).
The phytochrome (light) effect was supported by Yamaguchi et al. (1998), who showed that one of two 3-βhydroxylase enzymes, encoded by the GA4H gene is induced in germinating seeds by phytochrome. The mechanism of the GA signalling proteins and the effect of light is now well established. Crucial in this are the DELLA proteins which repress GA action, RGL2 (repressor of GA1-3 like 2) is the major DELLA regulating seed germination. These DELLA proteins are degraded by the 26S proteosome therefore GA charged GID1 DELLA proteins interact with the F box protein SLY1 needed for DELLA ubiquitination (Steber, 2007, Finkelstein et al., 2008). An important signal transduction component of light induced germination is the bHLH transcription factor PIF1 (Phytochrome Interacting Factor 1) also named PIL5 (Phytochrome-Interacting factor 3-Like 5) (Oh et al., 2004, 2006, Castillon et al., 2007), which can bind to Pfr that thereafter results in proteosome degradation (Oh et al., 2006). This increases GA levels because GA biosynthesis genes such as GA3ox1 and GA3ox2 are repressed by PIF1/PIL5, whereas the GA catabolic GA2ox gene and genes encoding DELLA proteins are activated (Oh et al., 2007). These higher GA levels further lead to inactivation of the DELLA proteins as described above. Somnus (SOM) which encodes a nucleus-localized CCCH-type zinc finger protein is another gene acting downstream of PIL5 (Kim et al., 2008). The som mutant germinates in darkness, independently of various light regimes. Kim et al. (2008) showed that PIL5 activates the expression of SOM by binding directly to the SOM promoter. It is suggested that PIL5 regulates ABA and GA metabolic genes partly through SOM.
Cold treatments were also found to stimulate GA biosynthesis (Yamauchi et al., 2004) by inducing the GA3ox1 and GA3ox2 genes. This cold effect is mediated by a light stable bHLH transcription factor SPATULA (SPT), which suppresses the expression of these genes (Penfield et al., 2005). In addition cold may increase the sensitivity of seeds to GAs because it also has an effect in GA deficient mutants (Debeaujon and Koornneef, 2000). The fact that often stratification is more effective than GA treatment (Alonso-Blanco et al., 2003) suggests that also other factors, promoting germination, are affected by stratification.
Brassinosteroids (BRs), are naturally occurring plant steroid hormones found in a wide variety of plant species (Clouse and Sasse, 1998) are also involved in the control of germination in Arabidopsis. The BR signal leads to reduced sensitive to ABA and thereby stimulates germination, although the normal germination of BR deficient and BR signalling mutants indicates that there is no absolute BR requirement for germination (Steber and McCourt, 2001). BRs could overcome the lack of germination of the sleepy1 mutant, probably by bypassing its GA requirement through a GA independent mechanism (Steber and McCourt, 2001; Finkelstein et al., 2008).
Mutants in ethylene signalling are also affected in their germination response. Ethylene is produced in trace amounts by almost all higher plants and is involved in the control of growth and developmental processes that range from germination to senescence. Often seeds that respond to ethylene are light sensitive for germination (Kepczynski and Kepczynska, 1997). Ethylene insensitive mutants such as etr and ein2 germinate less well or after a longer period of after-ripening than wild type (Bleecker et al., 1988; Beaudoin et al., 2000). The ein2 and etr mutants are hypersensitive to ABA (Beaudoin et al., 2000; Ghassemian et al., 2000), which agrees with the observation that ein2 mutants were isolated as abi1-1 suppressors. The ctr1 mutant, which is characterised by a constitutive ethylene response, appeared among mutants selected as enhancers of the ABA insensitive mutant abi1-1 and ctr1 monogenic mutants are also slightly ABA resistant (Beaudoin et al., 2000). These observations, in combination with the non-dormant phenotype of the ein2 abi3-4 double mutant indicated that ethylene negatively regulates seed dormancy by inhibiting ABA action (Beaudoin et al., 2000). However, Ghassemian et al., (2000) and Chiwocha et al. (2005) showed that, in addition to signalling, the slightly more dormant ethylene insensitive mutants, such as ein2 and etr1-2 have higher ABA levels. The higher levels of germination promoting hormones indicate compensation effects. The presence of cross talk between sugar signalling and ethylene was suggested by the sugar insensitive phenotype of ctr1 (Gibson et al., 2001) and the sugar hypersensitive phenotype of etr (Zhou et al., 1998). Apparently ABA, ethylene and sugar signalling strongly interact at the level of germination and early seedling growth.
Auxins are known to play important roles in embryogenesis. However, its role in the regulation of germination and seedling establishment remained obscure (Kucera et al., 2006). Auxin alone was not generally considered to be important in the control of seed germination but cross-talk between auxin, ABA, GA and ethylene was suggested to both affect germination and seedling establishment (Fu and Harberd, 2003; Ogawa et al., 2003; Chiwocha et al., 2005, Carrera et al., 2008; Liu, PP et al., 2007 reviewed by Holdsworth et al., 2008a). Analysis of the expression of the DR5:GUS auxin reporter indicated that auxin accumulates during embryogenesis and is present in the seed following imbibition. Expression was observed at the radicle tip prior to germination in one study (Liu, PP et al., 2007) and throughout the embryo at the end of embryogenesis in another study (Ni et al., 2001). Analysis of transcriptome expression showed that RNA encoding auxin transporters AUX1, PIN2, and PIN7 were highly up-regulated in response to treatment of ga1 mutant seeds with GA (Ogawa et al., 2003), and that both efflux and influx transporters are up-regulated in after-ripened compared to dormant seeds (Carrera et al., 2008). This may indicate a role for these transporters in germination per-se, or with the establishment of the root apex and gravitropism following radicle emergence. Clearer genetic evidence of a role for auxin in germination has been obtained from an analysis of the regulation of Auxin Response Factor10 (ARF10) by microRNA (miRNA) miR160 (Liu, PP et al., 2007). miRNAs have been shown to down-regulate target genes at the post-transcriptional level, and play crucial roles in a broad range of developmental processes (Dugas & Bartel, 2004). It was shown that transgenic seeds expressing an miR160-resistant form of ARF10 (mARF10) were hypersensitive to germination inhibition by exogenous ABA, whereas ectopic expression of miR160 resulted in reduced sensitivity to ABA (Liu, PP et al., 2007).
These results indicate a role of auxin in germination associated pathways, and suggest that interactions between auxin and ABA signalling pathways may contribute to the germination potential of seeds. An analysis of the function of key components of auxin signalling in relation to after-ripening, germination potential and vigour may reveal novel roles for auxin in these processes.
Compounds that have been identified as being important stimulants of germination, mainly using farmacological tools are several nitrogen-containing compounds, including nitric oxide (NO) gas (Bethke et al., 2006), nitrite (NO2−) and nitrate (NO3−) (Alboresi et al., 2005; Bethke et al., 2007a for review). It is suggested (Bethke et al., 2007a) that all N compounds affect germination via conversion into NO. Enzymatic NO production occurs mainly via nitrate reductase as by product of lipid catabolism or nitric oxide synthase (Crawford and Guo, 2005). Non-enzymatic conversion of nitrite to NO, has also been demonstrated and was suggested to have special significance for seeds (Bethke et al., 2007a). The observation by Alboresi et al. (2005) that the high nitrate levels that accumulate in nitrate reductase deficient mutants lead to reduced dormancy implies that nitrate reductase is not essential either because NO is generated by this non-enzymatic pathway or nitrate acts on its own.
An effect of NO is that it might function as an antioxidant (Lamatinna et al., 2003). However, it is also reported that NO inhibits catalase leading to higher H2O2 and reactive oxygen species (ROS), which has the opposite effect as antioxidants (Bethke et al., 2007a). ROS are a by product of β–oxidation of stored seed fatty acids. Thereby ROS are increased, which it self may alleviate dormancy (Bailly et al., 2004). However, other and additional mechanisms have also been suggested and include interaction with ABA catabolism enzymes and light and GA signalling (Finkelstein et al., 2008, Bethke et al., 2007b)