1. Which Turns Green in a Dark Box: Bean Sprouts or Pine Sprouts?
If you grow bean seeds in a box that lets no light in, the sprouts become yellowish-white and stretch out long. This happens because they cannot produce chlorophyll without light. Pines differ in this respect. Some gymnosperms, such as pine seedlings, are described as being able to green even in dark places. For flowering plants like peas, light is required to turn green [1]. It has also been reported that the cotyledons (the seed leaves) of Japanese black pines produce chlorophyll during germination in complete darkness [2].
It is important to note that this does not mean "the same amount as under light." Research reports show that cotyledons grown in the dark had fewer protein complexes related to photosynthesis than those grown in the light. They can become green even in the dark, but they are not always the same as plants in bright places [1]. So, how do they make chlorophyll without light?
2. How Beans and Pines Use Different Tools for the Last Step of Making Chlorophyll
Near the end of the pathway for making chlorophyll, there is a step where a substance called protochlorophyllide is changed into chlorophyllide a. Chlorophyllide a is the substance just before chlorophyll a. This change requires a reaction called reduction, and enzymes do this work [1].
There are two types of enzymes for this step. One is LPOR, which works using light. The other is DPOR, which does not use light and works in dark places. It is known that flowering plants only use LPOR to reduce protochlorophyllide. Because of this, protochlorophyllide builds up in the dark, forming structures called etioplasts, which are dark-grown precursors of chloroplasts [3]. Bean sprouts stay white because this step stops while waiting for light.
Gymnosperms that can green in the dark can also use DPOR. The parts of DPOR are made from three genes (chlL, chlN, and chlB) found in the DNA of the chloroplast. Gymnosperm chloroplast genomes have these three genes. Many flowering plants do not have at least two of them [4]. Whether or not they "have the tool that does not use light" is one reason pines and beans differ.
3. Do Pine Needles That Grow After the Cotyledons Also Turn Green in the Dark?
The discussion so far has mostly been based on studies of cotyledons. What about the early needles that grow after the cotyledons? A study from 1998 showed that early pine needles can also make chlorophyll in the dark. The two species examined were Pinus jeffreyi and Pinus sylvestris (Scots pine). Previous studies focused mainly on cotyledons, and it was not certain about needles [4].
In another study, the expression of DPOR genes was consistent in all samples analyzed for dark-grown seedlings of Pinus mugo and Pinus sylvestris. Proteins needed for chlorophyll synthesis, such as glutamyl-tRNA reductase, and proteins related to photosystem II, such as D1 and LHCI, were reported to accumulate regardless of light [5]. Even in the dark, the preparation for making green color proceeds steadily.
4. How Species and Temperature Change Pine Greening in the Dark
If you remember that "pines turn green even in the dark," you need to add conditions. According to reports, Japanese black pine cotyledons become yellow in very low temperatures in the dark, and light-independent chlorophyll synthesis is mostly suppressed. Also, there are differences in the strength of dark greening among pine family members; it is reported to be strong in Norway spruce and weak in larch [1]. Furthermore, there are examples of trees that have genes similar to DPOR but do not turn green in the dark.
Even within pine relatives, the degree of greening changes depending on species and temperature. It seems that just having the tool or not does not determine how green they become in the dark. So, where did this "tool that does not use light" come from?
5. How the Light-Free Enzyme DPOR Is Similar to Nitrogenase, an Enzyme That Fixes Nitrogen
DPOR is very similar to an enzyme called nitrogenase. Nitrogenase is the enzyme responsible for nitrogen fixation, which fixes nitrogen into a form living things can use. Articles introducing DPOR point out that there may be an evolutionary connection between nitrogen fixation and chlorophyll formation in the dark [1].
The enzyme that makes plant green color and the enzyme that grabs nitrogen might be relatives. Thinking about this, the roots of the tools inside the same green leaf might connect to unexpected places. However, being similar is not proof that they are relatives. It is only a possibility at this stage [1].
6. How to Test Beans and Pines at Home, and What to Read in Original Sources
You can check how bean sprouts change with or without light at home. Place bean seeds or mung bean sprout seeds on wet kitchen paper and divide them into two containers. Put one near a bright window and cover the other with a lightproof box to make it dark. Observe for about 3 or 4 days and compare the color and height. The sprouts in the box should often be whitish and stretch out long. After that, if you remove the box and place them by the window, you can also see changes in color.
If you want to investigate more, try searching for terms like DPOR, dark greening, and LPOR. Specific numbers about differences by pine species and temperature might be found in original research papers or review articles.