CELL-WALL LIGNIN CONTENT RELATED TO TRACHEID DIMENSIONS IN DROUGHT-SENSITIVE AUSTRIAN PINE (PINUS NIGRA) - Brill
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IAWA Journal, Vol. 22 (2), 2001: 113–120
CELL-WALL LIGNIN CONTENT RELATED TO TRACHEID DIMENSIONS
IN DROUGHT-SENSITIVE AUSTRIAN PINE (PINUS NIGRA)
by
Wolfgang Gindl
Institut für Botanik, Universität für Bodenkultur, Gregor Mendel Straße 33, A-1180 Wien, Austria
SUMMARY
The intra-annual distribution of cell-wall lignin concentration was de-
termined in Austrian pine tree rings and compared with tracheid diam-
eter, lumen width, cell wall thickness and proportion of cell wall area.
Lignin concentration was highly correlated with all tracheid dimensions,
but only the proportion of cell wall area exhibited a direct statistically
significant relationship. Since cell dimensions in Austrian pine are
subjected to the indirect and direct influences of the water status of
trees, the negative correlation between cellular lignin content and the
proportion of cell wall area is attributed to an indirect effect of water
stress on lignification in pine tracheids.
Key words: Lignin concentration, cell wall area, Pinus nigra Arnold,
water stress.
INTRODUCTION
Wood formation comprises the processes of cell division, cell enlargement, cell wall
thickening and lignification (Skene 1969; Wodzicki 1971). During the synthesis of
the secondary cell wall, deposition of carbohydrate polymers precedes lignification
(Kutscha & Schwarzmann 1975; Imagawa et al. 1976). The intra-annual variability
of tracheid characteristics in conifers is substantial and has been the subject of a number
of studies. Variations of tracheid dimensions have been analyzed thoroughly (e.g.,
Bannan 1964; Fengel & Stoll 1973; Grozdits & Ifju 1984; Vysotskaya & Vaganov
1989; Fujiwara & Iwagami 1990; Von Wilpert 1991), and the pattern of lignin distri-
bution in secondary cell walls across an annual increment is also known from a few
investigations (Fergus et al. 1969; Fukazawa & Imagawa 1981; Takano et al. 1983).
The highest lignin concentrations generally occur in the first earlywood cells fol-
lowed by a linear decrease towards the end of the growth ring. In the terminal late-
wood, variation increases and higher lignin content occasionally occurs. A part of the
variability of lignin concentration in terminal latewood may be explained by the influ-
ence of climatic extremes (Gindl & Grabner 2000).
The Austrian pine trees (Pinus nigra Arnold) selected for the present study are
characterized by a high sensitivity of radial growth to summer rainfall (Strumia et al.
1997). Furthermore, intra-annual density fluctuations frequently occur in years with
May drought events (Wimmer & Strumia 1998). The wide range of evenly distributed
tracheid sizes and cell wall thicknesses formed due to periodically occurring water
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stress make this tree species highly suitable for an investigation of relationships be-
tween tracheid dimensions and cellular lignin content as well as possible indirect
effects of the environment on lignin content.
MATERIAL AND METHODS
Increment cores from five Pinus nigra trees at Weikersdorf in the Vienna basin sam-
pled by Strumia et al. (1997) were selected for this study. The 1992 growth ring of
each increment core, characterized by the presence of intra-annual density fluctua-
tions (Wimmer & Strumia 1998), was identified by cross-dating with the Pinus nigra
master chronology built by Strumia et al. (1997). This growth ring was dissected from
the cores and embedded in Spurrʼs resin (Spurr 1969) after extraction and dehydra-
tion in ethanol and acetone. Cross sections (1 µm) were made on a Leica ultramicro-
tome equipped with a diamond knife. Sections were transferred to quartz slides and
observed at a × 1000 magnification (Zeiss MPM 800). Absorbance of ultraviolet light
in the middle of the secondary cell wall was determined at a wavelength of 280 nm
using a circular measuring spot with a diameter of 0.5 µm. At this small spot size
(Fig. 1), biasing effects of the highly lignified middle lamella on absorbance readings
in thin cell walls can be excluded. When measurements from more than one radial
cell file are averaged, the information associated with individual tracheids is lost,
even when tracheidogram procedures are applied (Vaganov 1990). Therefore, only
one cell file per tree was measured, averaging four determinations of absorbance per
tracheid. Lignin concentration was calculated from absorbance measurements accord-
ing to Scott et al. (1969).
After UV microscopy, the thin sections were stained with gentian violet and dig-
ital images were captured using a video camera mounted on a light microscope and
Fig. 1. Circular measuring spot (diameter = 0.5 µm) positioned in the middle of the secondary
cell wall of an Austrian pine tracheid. — Scale bar = 5 µm.
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analyzed with the software NIH-Image. Lumen width (LW) and cell wall thickness
(CWT) in radial direction were measured. Tracheid diameter was calculated by add-
ing 2 × CWT to LW. To estimate the proportion of cell wall area in the cross sectional
area of individual tracheids, a rectangular cross section and a constant tracheid diam-
eter of 35 µm in tangential direction were assumed.
RESULTS
Highly significant correlations between all investigated tracheid dimensions and lignin
content of the secondary cell wall (lignin per mass unit cell wall material) were found
using rank correlation analysis (Table 1). The relationship between lumen width and
lignin concentration is positive, i.e., the larger the lumen – the higher the lignin con-
centration, and best fitted by a logarithmic regression line, which is also the case for
the relationship between tracheid diameter and lignin concentration (Fig. 2). A nega-
tive correlation best fitted by a linear regression line was found for cell wall thickness
and lignin concentration (Fig. 3). The proportion of cell wall area achieves the high-
est negative correlation coefficient with lignin concentration. Like cell wall thick-
0.30
Lignin concentration (g g -1)
0.25
0.20
0.15
0.10
0 10 20 30 40 50
Lumen width (μm)
0.30
Lignin concentration (g g -1)
0.25
0.20
0.15
0.10
0 20 40 60
Tracheid diameter (μm)
Fig. 2. Plots of lignin concentration related to radial lumen width and radial tracheid diameter.
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Table 1. Spearman correlation coefficients between lignin concentration and tracheid dimen-
sions of Black pine. Significance of all correlations is p < 0.001 (n = 205).
Lumen Cell wall Tracheid Proportion of
width thickness width cell wall area
Lignin concentration 0.78 –0.68 0.63 –0.80
0.30
Lignin concentration (g g -1)
0.25
0.20
0.15
0.10
0 2 4 6 8 10
Cell wall thickness (μm)
0.30
Lignin concentration (g g -1)
0.25
0.20
0.15
0.10
0 0.2 0.4 0.6 0.8 1
Proportion of cell wall area
Fig. 3. Plots of lignin concentration related to radial cell wall thickness and proportion of cell
wall area.
ness, the proportion of cell wall area seems to be negatively and linearly correlated
to lignin concentration (Fig. 3). When partial correlations between lignin content
and either lumen width, tracheid diameter, cell wall thickness or proportion of cell
wall area are alternately calculated, controlling for the influence of the remaining
parameters, only the negative correlation between proportion of cell wall area and
lignin concentration remains significant (rpartial = -0.35, p < 0.001), indicating a direct
relationship between these two tracheid properties. Figure 4 shows an example of the
similarity of the intra-annual distribution of proportion of cell wall area and cell-wall
lignin content, illustrating the close negative relationship between these parameters.
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via free access1.0 0.15
Proportion of cell wall area
Lignin concentration
0.8
0.20
0.6
0.4
Gindl — Lignin content and tracheid dimensions
0.25
Lignin concentration (g g-1 )
Proportion of cell wall area
0.2
0.0 0.30
0 200 400 600 800 1000 1200
Distance from end of previous increment (μm)
Fig. 4. Negative relationship between the intra-annual distribution of cell-wall lignin concentration and proportion of cell wall area. Lignin concen-
tration is displayed on a reversed y-axis. The measurements were done in the central cell file of the displayed microsection.
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DISCUSSION
Concluding from the statistical analyses performed on data from 205 tracheids, cell-
wall lignin content is related to all four investigated tracheid dimensions, the relation-
ship between the proportion of cell wall area in an individual tracheid and the lignin
content of its secondary cell wall being the most robust. The variability of tracheid
dimensions in conifers is subjected to the indirect and direct effects of tree water
status (Zahner 1963). As long as an active apical meristem is present, water stress in
the tree crown has an indirect negative effect on the radial tracheid diameter mediated
by the concentration of auxin in the cambial zone (Larson 1963). After the cessation
of elongation growth, auxin concentration decreases, which is supposed to initiate the
transition from earlywood to latewood (Larson 1960). Indeed, the latter was found to
be highly correlated to the date of cessation of shoot elongation in Loblolly pine
(Jayawickrama et al. 1997). A direct effect of water stress is superimposed on this
relationship. Cell turgor associated with water supply is directly responsible for cell
enlargement (Kramer 1964; Von Wilpert 1991; Okuyama et al. 1995). Periods of pro-
nounced water stress initiate the formation of intra-annual density fluctuations (false
rings) (Zahner 1963), which are nothing else but fluctuations of the proportion of cell
wall area (Park & Telewski 1993), in the earlywood and latewood zones of conifers
(Kuo & McGinnes 1973).
Since the variability of cell dimensions in the investigated samples is so closely
related to water status, it seems reasonable to assume an indirect effect on cellular
lignin concentration. However, the effect of water stress on cell wall synthesis in
conifers is unclear. While Zahner (1963) attempts to explain the thick cell walls in
drought-induced density fluctuations by a relative increase in the availability of car-
bohydrates due to the reduced activity of the cambium, Whitmore and Zahner (1967)
found evidence for a direct negative influence of water stress on cell wall metabo-
lism. The finding that cell wall thickness and proportion of cell wall area are nega-
tively related to lignin content, whereas this relationship is positive for lumen width
and tracheid diameter, suggests a tradeoff between wall thickening (i.e. deposition of
carbohydrate polymers) and lignification due to the limited capacity of the protoplast
to synthesize cell wall components.
In the present study, it could be demonstrated that cellular lignin content in drought-
stressed Austrian pine is highly correlated with tracheid dimensions, and an indirect
effect of tree water status on lignin content is proposed. The physiological mecha-
nism relating lignification to cell dimensions remains to be uncovered and will be the
subject of future research.
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