Leaf shedding and NP remobilization peaked in mid-summer, while growth peaked in spring
A number of previous works have reported a tight chronological dependence of growth events and internal nutrient remobilization (Fife & Nambiar, 1982; Mark & Chapin, 1989; Nambiar & Fife, 1991; Jaeger & Monson, 1992; Oliveira et al., 1996; Milla et al., 2004). Here, however, although the seasonal distribution of N and P remobilization overlapped with part of the phenological activities, a remarkable delay between the peak in vegetative growth and flowering, and the peaks in N and P remobilization, was detected. In addition, K remobilization was nearly unrelated to growth phenology.
The time-lag between the peak in vegetative growth and N and P remobilization seems to be driven by the calendar of leaf abscission. Most of our species shed their leaves after DVG, mainly at the beginning of the summer. Nutrients reabsorbed during leaf senescence may account for a large proportion of the nutrients remobilized along the whole year (Cherbuy et al., 2001). This contrasts with other evergreen species, where the remobilization of nutrients independently of leaf senescence is the major mechanism of N redistribution in the branches (Nambiar & Fife, 1991; Millard & Proe, 1993; Wendler et al., 1995; Pasche et al., 2002). By contrast, in the species of this study, leaf senescence appears to be the chief process leading to N and P remobilization. As pointed out in the Introduction section, spring–summer leaf senescence can be triggered by factors such as growth events or the arrival of summer drought in Mediterranean evergreens (Munné-Bosch & Alegre, 2004). Vegetative growth nutrient requirements had been fulfilled, either from soil uptake or from remobilization from roots or trunks, before the time when remobilization in the branches and leaf shedding peaked. The latter might therefore have been used to replenish nutrient reserves of older organs. However, leaf senescence might also have been promoted by the need of adjusting water transpiring surface at the beginning of the summer drought (Del Arco et al., 1991).
An alternative explanation to the delay between growth and remobilization could lie in the seasonality of soil nutrient supply. Vegetative growth is more vigorous and frequently overlaps with flowering at mid-spring. In this period, soil nutrient availability and nutrient uptake by roots are high (Bonilla & Rodá, 1992; Serrasoles et al., 1999), and could provide a large proportion of the nutrients required for growth. Both N and P remobilization peak sharply in mid-summer when soil supply is scarce. According to some authors, N remobilization predominates when environmental supply is low (Wendler et al., 1995; Silla & Escudero, 2003). The formation of overwintering buds begins immediately after the elongation of vegetative shoots (Yuceer et al., 2003), thus during the summer peak in N and P remobilization in the species of this study. In species with determinate growth, a large amount of N imported into the new organs occurs during bud growth (Millard, 1994). Furthermore, given that shoot extension slows down in summer, bud formation is likely to occur in a period of low external nutrient supply, and therefore requires more nutrients from internal remobilization. Therefore, although leaf senescence and massive N and P remobilization are unlikely to be triggered by the morphogenesis of overwintering buds, the latter probably benefited from the temporal coincidence of bud growth and N and P remobilization in the stressful Mediterranean summer. We should note that below-ground phenology was not monitored, and root growth might be an additional trigger of nutrient remobilization.
Nitrogen and P have some similar, but many distinct physiological functions. A large amount of N in green leaves is bound to the proteins of the Calvin cycle and thylakoids (Urban et al., 2004), which are nearly absent in nonphotosynthetic (or poorly photosynthetic) reproductive organs. On the other hand, there is a great demand of P during flowering (Ashman & Baker, 1992; Niva et al., 2003). In addition, P limits sexual reproduction in infertile environments (Nagy & Proctor, 1997; Brouwer et al., 2001). Accordingly, N : P ratios were systematically higher in the vegetative than in the reproductive organs of the species of this study (data not shown), as has been found elsewhere (Ågren, 1988; Karlsson, 1994; Güsewell, 2004). Despite such differences, the seasonal patterns of N and P remobilization were very similar. This might be due to two facts. First, most biomass and nutrient pools in the branches were in vegetative organs (see Fig. 3). Therefore, despite P being more related to reproductive functions than N, P remobilization was more dependent on the growth of vegetative sinks than on reproductive, because of the smaller size of the latter. Second, most N and P remobilization probably came from leaf senescence in the branches of this study (see Fig. 2). During leaf senescence, N and P resorption often correlate positively, irrespective of the relative needs of each nutrient in the plant (Killingbeck, 1996).
The remobilization of K from the old parts of the branch peaked in periods of the year that differed greatly in phenological stage of the plants and climate. The main K remobilization event occurred in mid-summer, as did N and P, and could be due to K resorption before leaf shedding, or to the fulfilment of nutrient demands when soil availability is low, as explained earlier. However, K plays a crucial role in stabilizing cell pH and in regulating osmotic potential (Lansac et al., 1994; Marschner et al., 1997). In recently born organs, osmotic adjustment during the summer is required to endure drought conditions (Sanchez-Blanco et al., 2002; Tognetti et al., 2002). In addition, under moderate water stress, the accumulation of osmolytes in growing organs helps to increase cell turgor, thus facilitating growth (Mengel & Arneke, 1982; Boyer, 1988). Therefore, the remobilization of K in the summer could contribute to the maintenance of growth during this period. The depletion of K that occurs in mid-winter may also be related to the osmotic properties of this element. In mid- to late winter the renewal buds of most of our species start to swell. During this period evapotranspiration (Savéet al., 1999) and temperatures are low. Given that K drives water to the meristems through osmotic water lifting (Zimmermann et al., 2002), and also accumulates in tissues subjected to cold stress (Fernandez et al., 2003), bud growth in winter could be facilitated by K remobilization from branch stores to the meristematic tips. Other studies also report that N is more related to growth requirements, and K to osmotic and carbohydrate transport functions (Proe et al., 2000). However, in our calculations of %NurBR(month) we did not consider leaching as a potential mechanism of nutrient loss between sampling dates. Given that leaf K is easily leachable from the foliage (Wang et al., 2003), this process could have accounted for some K losses during rainy months.
The amount of NPK remobilized per gram of branch was directly related to that accumulated in the growing vegetative organs along a season
If growth events were to exert a strong immediate effect on the remobilization of branch nutrient stores, this should be reflected in a close coupling between the outcome of both processes (nutrient pools in new organs, and remobilized nutrient pools from older organs) by the end of the growing season. In fact, branches that used more N, P or K for growth along the season also showed higher depletion of nutrient stores. Therefore, despite the relative asynchrony between growth and nutrient remobilization, the amount of reabsorbed nutrients correlated directly with the nutrient use for vegetative growth.
However, taking into account the time-lag between vegetative growth and remobilization, nutrients cannot be diverted directly from old to new vegetative biomass in the single branch, because remobilization occurs mainly after growth. What then is the rationale for this correlation? Several explanations can be suggested in this regard, though the selection of one of them would be speculative in the light of our data. First, the expansion of a new cohort of leaves probably changes the hormonal balances within the branch. As a consequence, old leaves should become a weaker sink for metabolites, and therefore are likely to begin senescence. Second, provided that growth events in long-lived perennials are highly dependent on internal remobilization (Aerts & Chapin, 2000), nutrient reserves other than old leaves (i.e. trunks or roots) are probably depleted of nutrients in early spring, which might be later replenished by the remobilization of nutrients from senescing leaves. Such complex seasonal remobilization dynamics have been reported previously in Rhododendron (Pasche et al., 2002; Lamaze et al., 2003). Moreover, in a number conifer species, a portion of the pool of nutrients taken up by roots and reabsorbed during the senescence of old needles in a given season is also temporarily stored in woody and needle tissues and invested later in the new vegetative growth during the following spring. This results in a close coupling between previous-year nutrient stores, and current-year nutrient use in growth, irrespective of current-year nutrient supply from the environment (Millard & Proe, 1993; Proe & Millard, 1994; Millard, 1996; Proe et al., 2000). Third, the plant can supply water to a limited number of leaves during summer, and therefore the larger the newly expanded leaf area the higher the number of old leaves which have to be shed. This would imply that the measured relation between nutrient use and supply is an indirect effect of the adjustment of leaf area when shoot growth has finished and summer drought is about to arrive. Previous work on the significance of nutrient remobilization for growth has rarely considered the chronology of both events. Our findings suggest that simple correlations between investments and supply can be incorrectly interpreted as direct source–sink functional relationships, if we lack phenological data.
Previous research on perennial or biennial herbs and grasses emphasized the dependence of reproductive events on internal N and P cycling (Heilmeier et al., 1986; Mark & Chapin, 1989; Jaeger & Monson, 1992; Bausenwein et al., 2001). In our study, although reproductive growth and N and P remobilization peaked in summer or at the end of spring, and therefore nutrient remobilization might be important to supply nutrients for reproductive events, the amount of nutrients remobilized was unrelated to the needs of reproductive organs. This was probably because of the comparatively lower resource use of reproductive growth (see Fig. 3) and, in accordance with the explanation in the preceding paragraph, to the fact that the development of reproductive structures does not display new leaf area which might trigger senescence-related nutrient remobilization.
In summary, in the branches of the studied species, N and P remobilization depend more on the chronology of leaf shedding than on growth events. Provided that the mineral nutrition of adult individuals of long-lived perennials is highly dependent on remobilization (Aerts & Chapin, 2000), the studied species should make use of organs other than the leafy branches which act as an intermediate storage site for the amounts of nutrients remobilized in summer and those used in the following spring growth event. The shortage of soil nutrient supply during summer drought could also promote nutrient remobilization in summer. Potassium remobilization appears to depend on factors such as demands for osmotic adjustment in climatically harsh periods of the year.