Reply on RC1

REPLY: We opted for Biogeosciences instead of a plant physiology journal because we envisage a more general audience for our results. Although the measurements presented in the manuscript focus for a large part on leaf-level responses, the main incentive for this study was the importance of nutrient availability and plant-mycorrhiza interactions in determining carbon cycling. We expect that our results will be useful for the land surface modeling community and particularly to those who are aiming to implement carbonnutrient interactions in these models.

made in the manuscript. The line numbers (L_ _) in our replies refer to those of the original manuscript.
1) I was a bit surprised that this manuscript was submitted to Biogeosciences, as I would have thought that a straight physiology or ecophysiology journal would have been more appropriate. However, I will leave that aspect of my feedback to the Editor.

REPLY:
We opted for Biogeosciences instead of a plant physiology journal because we envisage a more general audience for our results. Although the measurements presented in the manuscript focus for a large part on leaf-level responses, the main incentive for this study was the importance of nutrient availability and plant-mycorrhiza interactions in determining carbon cycling. We expect that our results will be useful for the land surface modeling community and particularly to those who are aiming to implement carbonnutrient interactions in these models.
2) The authors quickly jumped to the conclusion that P effects must all be direct effects on photosynthesis, and appear rather dismissive of the idea that P might effect leaf growth, and the change in sink demand would then affect photosynthetic activity. Even though they cite some of the papers highlighting P effects on sink activity, with effects of photosynthesis being indirect, the message in those papers appears not to have been taken on board. Tools exist to assess feedback inhibition of photosynthesis, but the literature dealing with that aspect wasn't discussed at all. For example: Sharkey T D, Stitt M, Heineke D, Gerhardt R, Raschke K and Heldt H W 1986 Limitation of photosynthesis by carbon metabolism: II. O2-insensitive CO2 uptake results from limitation of triose phosphate utilization. Plant Physiol. 81, 1123Physiol. 81, -1129 In the revised manuscript, we will add extra text on the direct and indirect effects of P on photosynthesis, including the possibility for changes in sink demand: Inorganic phosphate (Pi) directly affects the activity of Calvin cycle enzymes through the level of activation. For instance, Pi is required for light activation of Rubisco (Parry et al., 2008). It also directly affects maximum rate of CO 2 -limited carboxylation (v cmax ) and triose phosphate utilization (Lewis et al., 1994) and RuBP-regeneration-limited rates of electron transport (Loustau et al., 1999). On the other hand, Pi can indirectly affect photosynthesis through the changes in stromal pH (Bhagwat 1981), where the consumption of Pi as a substrate of photosynthesis could decrease photosynthesis by a direct effect of low stromal Pi concentration on Rubisco. Moreover, the effect of P on photosynthesis depends on the dynamic interactions between sink and source tissues. The low P level decreases sink strength which imposes the primary limitation on photosynthesis (Pieters et al., 2001). Pi deprivation impacts on photosynthesis can also be explained by diminishing carbon export to sinks (Pieters et al., 2001). Moreover, low sink strength lowers sucrose synthesis and restricts the recycling of Pi back to the chloroplast thus limiting the rate of net photosynthesis (Paul and Foyer, 2001). Decreasing cytoplasmic Pi also limits the photosynthesis through end product (feedback) inhibition and this end-product inhibition could be due to high concentrations of triose-P. In this regard The total Pi within the chloroplast is relatively constant, thus high triose-P is automatically coupled with low Pi, which in turn could limit photosynthesis. For example, Pi deficiency drastically decreased RuBP content in the Pi-deficient leaves and hence the rate of photosynthesis. On the other hand, at high Pi supply, triose-P export competes with ribulose 1,5-bisphosphate (RuBP) regeneration and the rate of photosynthesis can be diminished.
Bhagwat, A. S.: Activation of spinach ribulose 1,5-bisphosphate carboxylase by inorganic phosphate. Plant Sci. Lett., 23, 197-206, 1981. 3) The authors need to consult a recent textbook to check where different reactions related to carbon metabolism in C4 plants occur, because it is not correct that synthesis of starch and sucrose occur in different cell types. Both require Rubisco, which only occurs in the bundle-sheat cells, and not in mesophyll cells. We consulted other sources, but found similar information.

REPLY
"Sucrose was predominantly synthesized in the mesophyll cells and starch in the bundle sheath cells" (Furbank and Kelly, 2021). "In Zea mays L. and Atriplex spongiosa F. Muell., sucrose-phosphate synthase (key enzyme in sucrose biosynthesis) was located almost exclusively in the mesophyll cells" (Lunn and Furbank, 1997). We can add these extra references to the manuscript, or if preferred, delete the phrases that are in doubt. 4) It is true that mycorrhizas may mobilise organic P or sorbed P, but when it comes to arbuscular mycorrhizas (AM), the cited textbook (Smith & Read) points out that AM are unlikely to do that. Their role is to enhance the volume of soil that can be explored. So, the text needs to be tweaked a bit to acknowledge that.

REPLY:
Here we disagree with the reviewer. Many studies have shown that AMF (contrary to ectomycorrhizae) are especially important for plant uptake of P, not only because of the increased soil volume explored, but because they produce exudates that liberate P from the minerals ( 5) SLA is not a simple measure of leaf thickness, but of both leaf thickness and leaf density. Leaf density is affected by carbohydrate concentrations and amount of cell walls.

REPLY:
The reviewer made a good point here. With the thinner feel of the leaves in mind, we went too short here. The higher SLA indeed points to lower leaf density and/or leaf thickness. The lower concentration of leaf compounds in the non-P-fertilized mesocosms (as shown in table 1) might suggest lower leaf densities indeed. We will adapt this in the manuscript. 6) 'Content' is generally used when amounts are expressed per plant (part); when amounts are expressed per unit mass or area, 'concentration' is recommended.

REPLY:
We agree with the reviewer. We will change 'content' to 'concentration' where appropriate.
Below we list the additional comments made by Referee 1 in the manuscript. 7) L15: Why start with 'Despite'? I suggest to start with 'Phosphorus' and wrote this as two statements about P. (Despite doesn't make sense here.)

REPLY:
We will change this in the revised manuscript to ' Phosphorus (P) is an essential macronutrient for plant growth and one of the least available nutrients in soil. P limitation is often a major constraint for plant growth globally.' 8)L17-18: Change '...experiments have been carried out to study the long-term effects on the yield, data on P addition effects to seasonal variation in leaf-level photosynthesis are scarce.' To '...experiments have been carried out to study the long-term effects on yield, data on P addition effects on seasonal variation of leaf-level photosynthesis are scarce.'

REPLY:
We will make this change as suggested. 9) L20-22: The primary effect of P is just as likely on growth, rather than photosynthesis, and effects on photosynthesis likely reflect a reduced sink demand on source activity.

REPLY:
See reply to comment 2 above. 10) L34: Replace 'participates in the formation' by 'is a component'.

REPLY:
We will adapt as suggested. 11) L35-36: I don't know what this means, but do know that P is important in Pcontaining metabolites that play a role in carbon metabolism. Schulze et al. is an odd reference here. I think this one would be more appropriate: Veneklaas E J, Lambers H, Bragg J, Finnegan P M, Lovelock C E, Plaxton W C, Price C, Scheible W-R, Shane M W, White P J and Raven J A 2012 Opportunities for improving phosphorus-use efficiency in crop plants. New Phytol. 195, 306-320. 10.1111/j.1469-8137.2012.04190.x.

REPLY:
We will add the suggested reference. 12) L38: I can see how plants experience P stress, or, rather plant productivity is limited by P, but lands doesn't really experience P stress.

REPLY:
We will rephrase this sentence to 'On more than one third of the arable land worldwide, plant productivity is considered to be limited by P.' 13) L40: Delete 'the' in '…effect on the yield'.

REPLY:
We will delete as suggested.

REPLY:
We thank the reviewer for this suggestions and will add these references to the manuscript. 16) L47 'P is required for adenosine triphosphate (ATP) synthesis': This is true, but ATP is only a minute fraction of the metabolite P pool. See: Veneklaas E J, Lambers H, Bragg J, Finnegan P M, Lovelock C E, Plaxton W C, Price C, Scheible W-R, Shane M W, White P J and Raven J A 2012 Opportunities for improving phosphorus-use efficiency in crop plants. New Phytol. 195, 306-320. 10.1111/j.1469-8137.2012.04190.x.

REPLY:
We will add the reference to the manuscript. 17) L48 'P-deficiency therefore leads to a decrease in RuBP pool size and insufficient ATP, and consequently to a decrease in photosynthetic C assimilation.': Or is that decline due to a decreased sink demand?
REPLY: See reply to comment 2 above. 18) L53: Low sink demand for sugars, and feedback inhibition of photosynthesis: Sharkey T D, Stitt M, Heineke D, Gerhardt R, Raschke K and Heldt H W 1986 Limitation of photosynthesis by carbon metabolism: II. O2-insensitive CO2 uptake results from limitation of triose phosphate utilization. Plant Physiol. 81, 1123-1129.