The CBF1-dependent low temperature signalling pathway, regulon and increase in freeze tolerance are conserved in Populus spp.

MAPK (mitogen-activated protein kinase) pathways have been implicated in stress signalling in plants. In the present study, we performed yeast two-hybrid screening to identify partner MAPKs for OsMKK (Oryza sativa MAPK kinase) 6, a rice MAPK kinase, and revealed specific interactions of OsMKK6 with OsMPK3 and OsMPK6. OsMPK3 and OsMPK6 each co-immunoprecipitated OsMKK6, and both were directly phosphorylated by OsMKK6 in vitro. An MBP (myelin basic protein) kinase assay of the immunoprecipitation complex indicated that OsMPK3 and OsMPK6 were activated in response to a moderately low temperature (12°C), but not a severely low temperature (4°C) in rice seedlings. A constitutively active form of OsMKK6, OsMKK6DD, showed elevated phosphorylation activity against OsMPK3 and OsMPK6 in vitro. OsMPK3, but not OsMPK6, was constitutively activated in transgenic plants overexpressing OsMKK6DD, indicating that OsMPK3 is an in vivo target of OsMKK6. Enhanced chilling tolerance was observed in the transgenic plants overexpressing OsMKK6DD. Taken together, our data suggest that OsMKK6 and OsMPK3 constitute a moderately low-temperature signalling pathway and regulate cold stress tolerance in rice.

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Supercooling in Dormant Flower Buds of Forsythia, and the Correlation between Pistil Size and Bud Hardiness

Journal of Environmental Horticulture ◽

10.24266/0738-2898-17.2.57 ◽

1999 ◽

Vol 17 (2) ◽

pp. 57-62 ◽

Cited By ~ 1

Author(s):

Cindy L. Flinn ◽

Edward N. Ashworth

Keyword(s):

Low Temperature ◽

Supercooled Water ◽

Freeze Tolerance ◽

Freezing Injury ◽

Laboratory Studies ◽

Flower Buds ◽

Deep Supercooling ◽

Freeze Stress ◽

Stem Segments ◽

Accurate Indicator

Abstract Experiments were conducted to determine if dormant buds of Forsythia taxa exhibit the deep supercooling characteristic. Specimens were collected from thirteen Forsythia taxa including: F. suspensa (Thunb.) Vahl, F. x intermedia cv. Spectabilis (Koehne), F. x intermedia cv. Lynwood (G.E. Peterson), F. europaea (Degen and Baldacci), F. giraldiana (Lingelsh), F. japonica (Makino) var. saxatilis (Nakai), F. mandshurica (Uyeki), F. ovata (Nakai), F. suspensa var. fortunei (Lindl.), F. viridissima (Lindl.), F. x intermedia cv. Arnold Giant (Sax), F. cv. Arnold's Dwarf, and F. cv. Meadowlark (Flint). Buds and attached stem segments, were cooled at 2C (3.6F) per hour, and the temperature at which freezing occurred was determined by thermal analysis. Typically, two distinct freezing events were detected within Forsythia buds. The first freezing event, or high temperature exotherm, occurred just below 0C (32F), while the second freezing event, or low temperature exotherm, occurred between −16C (3.2F) and −28C (−18.4F). The low temperature exotherm corresponded to the freezing of supercooled water within dormant buds, and the detection of low temperature exotherms in buds of all 13 Forsythia taxa indicated that deep supercooling is common among members of this genus. In nine of the 13 Forsythia taxa, the temperature of the low temperature exotherm was an accurate indicator of bud freeze-tolerance (LT50), as determined by a laboratory freeze-stress protocol. The discrepancies noted in the other four taxa were apparently due to the occurrence of field freezing injury prior to conducting these laboratory studies. Evidence indicated a relationship between the extent of supercooling and the size of the pistil in dormant Forsythia buds.

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Changes in Freezing Tolerance, Water Potential, and Gene Expression of Poncirus trifoliata `Rubidoux' Seedlings Exposed to Acclimating Low Temperatures and Long Days

HortScience ◽

10.21273/hortsci.31.4.580a ◽

1996 ◽

Vol 31 (4) ◽

pp. 580a-580

Author(s):

Milton E. Tignor ◽

John M. Davis ◽

Frederick S. Davies ◽

Wayne B. Sherman

Keyword(s):

Gene Expression ◽

Low Temperature ◽

Water Potential ◽

Freezing Tolerance ◽

Freeze Tolerance ◽

Poncirus Trifoliata ◽

Economic Losses ◽

Tissue Samples ◽

Coumarate:Coa Ligase ◽

Hardy Cross

Poncirus trifoliata is a comparatively hardy, cross compatible, and graft compliant relative of Citrus. The citrus industry in Florida has suffered immense economic losses due to freezes. Although much research has been done in citrus freeze hardiness, little work has been on the early induction of freeze tolerance by low temperature. Poncirus trifoliata `Rubidoux' seedlings were germinated in perlite under intermittent mist at about 25°C and natural daylight conditions in a greenhouse and grown 2 weeks. See dlings were then transferred into a growth chamber at 25°C and 16 hour daylength for 1 week. Temperature was lowered to 10°C and tissue samples were collected at 0, 6, 24, and 168 hours. Freezing tolerance, at –6.7°C as determined by electrolyte leakage, and stem (leaves attached) water potential, measured using a pressure bomb, were also recorded for a subset of seedlings for the above intervals. After exposure to low temperature for 48 hours a red coloration became visible at the petiole leaflet junction an d at the buds, with subsequent exposure to low temperature the coloration spread to the leaves. Clones for phenylalanine ammonia lyase (PAL), 4-coumarate:CoA ligase (4CL), and chlorophyll ab binding protein (CAB), and chalcone synthase (CHS) were used to probe RNA isolated from P. trifoliata. PAL and 4CL transcripts increased in response to the low temperature. Significant increases in freeze hardiness occurred within 6 hours in the leaves, and increases continued for up to one week. Water potential increased from –0.6 to –2.0 MPa after 6 hours, then returned to –0.6 MPa after 1 week. These data indicate that increases in freezing tolerance and changes in water potential and gene expression can be detected shortly after low temperature treatments are imposed on P. trifoliata seedlings.

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Low-temperature Exotherms and Freeze Tolerance in Three Taxa of Deciduous Trees

HortScience ◽

10.21273/hortsci.30.4.849a ◽

1995 ◽

Vol 30 (4) ◽

pp. 849A-849

Author(s):

Orville M. Lindstrom ◽

Tomasz Anisko ◽

Michael A. Dirr

Keyword(s):

Low Temperature ◽

Cold Hardiness ◽

Acer Rubrum ◽

Freeze Tolerance ◽

Reliable Estimate ◽

Fraxinus Americana ◽

Deep Supercooling ◽

The Difference ◽

The Relationship ◽

Zelkova Serrata

Although differential thermal analysis has been routinely used to evaluate cold hardiness, the relationship between deep supercooling ability and plant survival is not clear. We compared seasonal profiles of changes in low-temperature exotherm (LTE) occurrence and visually determined lowest survival temperature (LST) of Acer rubrum `Armstrong', Fraxinus americana `Autumn Purple' and Zelkova serrata `Green Village' growing in three locations representing plant cold hardiness zones 8, 7 and 5. Between December and February, LTE in Acer rubrum and Fraxinus americana occurred at temperatures 10 to 25C lower than the LST. The difference between LTE and LST was not significant for Zelkova serrata from January to April, and for Acer rubrum and Fraxinus americana in March. Data indicate that LTE could be used as an estimate of LST in Zelkova serrata but not in Acer rubrum and Fraxinus americana. This study demonstrated that LTE does not provide a reliable estimate of cold hardiness in all species that deep supercool.

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Freeze tolerance: constraining forces, adaptive mechanisms

Canadian Journal of Zoology ◽

10.1139/z88-164 ◽

1988 ◽

Vol 66 (5) ◽

pp. 1122-1127 ◽

Cited By ~ 12

Author(s):

Kenneth B. Storey ◽

Janet M. Storey

Keyword(s):

Protein Structure ◽

Low Temperature ◽

Cell Volume ◽

Thermal Hysteresis ◽

Freeze Tolerance ◽

Cell Integrity ◽

Physical Damage ◽

Term Survival ◽

Subzero Temperatures

For a variety of ectothermic animals, survival of subzero temperatures is aided by a natural capacity to tolerate extracellular freezing. Both low temperatures and freezing place inescapable constraints on the behaviour of molecules and biological structures. For example, low temperature affects metabolic rates, membrane fluidity, and weak bond interactions governing protein structure and function, whereas damage from freezing includes osmotic stress, membrane deformation, dehydration, physical damage by ice, and the consequences of long-term ischaemia. Selected biochemical adaptations permit survival of exposure to freezing by maintaining cell integrity, subcellular structure, energy production, and homeostasis. The key adaptations for freeze tolerance deal with the following: (i) control of extracellular ice: ice-nucleating proteins induce ice formation at multiple extracellular sites and at high subzero temperatures, whereas thermal hysteresis proteins inhibit the recrystallization of ice during long-term freezing; (ii) regulation of cell volume: the colligative action of high concentrations of polyols limit freeze concentration of the cell beyond a critical cell volume; (iii) protection of subcellular organization: trehalose and proline stabilize membrane bilayer structure, polyols stabilize protein structure; and (iv) viability in the frozen state: a well-developed tolerance for ischaemia plus mechanisms of facultative metabolic depression support long-term survival. Potential constraints of low temperature on metabolic functions are overcome to produce a metabolism that remains integrated and balanced over a wide temperature range. In addition, temperature change is exploited as a signal for the induction of various freeze tolerance adaptations.

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Exsolution and inversion in pigeonite from the Whin Sill, Northern England

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100109513 ◽

1978 ◽

Vol 36 (1) ◽

pp. 474-475

Author(s):

P.P.K. Smith

Keyword(s):

High Temperature ◽

Low Temperature ◽

Homogeneous Nucleation ◽

Silicate Mineral ◽

Antiphase Boundaries ◽

Two Phase ◽

Primitive Form ◽

The Core ◽

Nucleation Mechanisms ◽

And Inversion

Grains of pigeonite, a calcium-poor silicate mineral of the pyroxene group, from the Whin Sill dolerite have been ion-thinned and examined by TEM. The pigeonite is strongly zoned chemically from the composition Wo8En64FS28 in the core to Wo13En34FS53 at the rim. Two phase transformations have occurred during the cooling of this pigeonite:- exsolution of augite, a more calcic pyroxene, and inversion of the pigeonite from the high- temperature C face-centred form to the low-temperature primitive form, with the formation of antiphase boundaries (APB's). Different sequences of these exsolution and inversion reactions, together with different nucleation mechanisms of the augite, have created three distinct microstructures depending on the position in the grain.In the core of the grains small platelets of augite about 0.02μm thick have farmed parallel to the (001) plane (Fig. 1). These are thought to have exsolved by homogeneous nucleation. Subsequently the inversion of the pigeonite has led to the creation of APB's.

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Low temperature x-ray microanalysis of frozen-hydrated tobacco leaves

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100111537 ◽

1984 ◽

Vol 42 ◽

pp. 284-285

Author(s):

S. Edith Taylor ◽

Patrick Echlin ◽

May McKoon ◽

Thomas L. Hayes

Keyword(s):

Low Temperature ◽

Lemna Minor ◽

Root Tips ◽

Tobacco Leaves ◽

X Ray ◽

Whole Cells ◽

Carbon Coated ◽

The Right ◽

Beam Currents ◽

The University

Low temperature x-ray microanalysis (LTXM) of solid biological materials has been documented for Lemna minor L. root tips. This discussion will be limited to a demonstration of LTXM for measuring relative elemental distributions of P,S,Cl and K species within whole cells of tobacco leaves.Mature Wisconsin-38 tobacco was grown in the greenhouse at the University of California, Berkeley and picked daily from the mid-stalk position (leaf #9). The tissue was excised from the right of the mid rib and rapidly frozen in liquid nitrogen slush. It was then placed into an Amray biochamber and maintained at 103K. Fracture faces of the tissue were prepared and carbon-coated in the biochamber. The prepared sample was transferred from the biochamber to the Amray 1000A SEM equipped with a cold stage to maintain low temperatures at 103K. Analyses were performed using a tungsten source with accelerating voltages of 17.5 to 20 KV and beam currents from 1-2nA.

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Bulk standards for low-temperature biological x-ray microanalysis

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100111525 ◽

1984 ◽

Vol 42 ◽

pp. 282-283

Author(s):

P. Echlin ◽

M. McKoon ◽

E.S. Taylor ◽

C.E. Thomas ◽

K.L. Maloney ◽

...

Keyword(s):

Phase Separation ◽

Low Temperature ◽

Analytical Method ◽

Salt Solutions ◽

Absolute Concentration ◽

Cooling Process ◽

X Ray ◽

The Absolute ◽

Analytical Procedures ◽

Electrical Conductors

Although sections of frozen salt solutions have been used as standards for x-ray microanalysis, such solutions are less useful when analysed in the bulk form. They are poor thermal and electrical conductors and severe phase separation occurs during the cooling process. Following a suggestion by Whitecross et al we have made up a series of salt solutions containing a small amount of graphite to improve the sample conductivity. In addition, we have incorporated a polymer to ensure the formation of microcrystalline ice and a consequent homogenity of salt dispersion within the frozen matrix. The mixtures have been used to standardize the analytical procedures applied to frozen hydrated bulk specimens based on the peak/background analytical method and to measure the absolute concentration of elements in developing roots.

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Field ion microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100104236 ◽

1988 ◽

Vol 46 ◽

pp. 434-435

Author(s):

Gert Ehrlich

Keyword(s):

Low Temperature ◽

Sample Surface ◽

Low Pressure ◽

Radius Of Curvature ◽

Field Ion Microscopy ◽

Phosphor Screen ◽

Ion Microscopy ◽

Field Ion Microscope

The field ion microscope, devised by Erwin Muller in the 1950's, was the first instrument to depict the structure of surfaces in atomic detail. An FIM image of a (111) plane of tungsten (Fig.l) is typical of what can be done by this microscope: for this small plane, every atom, at a separation of 4.48Å from its neighbors in the plane, is revealed. The image of the plane is highly enlarged, as it is projected on a phosphor screen with a radius of curvature more than a million times that of the sample. Müller achieved the resolution necessary to reveal individual atoms by imaging with ions, accommodated to the object at a low temperature. The ions are created at the sample surface by ionization of an inert image gas (usually helium), present at a low pressure (< 1 mTorr). at fields on the order of 4V/Å.

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High-Vacuum Evaporation and a Cryostage to Observe Fractured Yeast

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100103346 ◽

1988 ◽

Vol 46 ◽

pp. 256-257

Author(s):

William P. Wergin ◽

Eric F. Erbe ◽

Eugene L. Vigil

Keyword(s):

Electron Microscope ◽

Low Temperature ◽

Standard Specimen ◽

High Vacuum ◽

Specimen Holder ◽

Sputter Coating ◽

Fresh Frozen ◽

Gain Access ◽

Scanning Electron ◽

Transfer Device

Investigators have long realized the potential advantages of using a low temperature (LT) stage to examine fresh, frozen specimens in a scanning electron microscope (SEM). However, long working distances (W.D.), thick sputter coatings and surface contamination have prevented LTSEM from achieving results comparable to those from TEM freeze etch. To improve results, we recently modified techniques that involve a Hitachi S570 SEM, an Emscope SP2000 Sputter Cryo System and a Denton freeze etch unit. Because investigators have frequently utilized the fractured E face of the plasmalemma of yeast, this tissue was selected as a standard for comparison in the present study.In place of a standard specimen holder, a modified rivet was used to achieve a shorter W.D. (1 to -2 mm) and to gain access to the upper detector. However, the additional height afforded by the rivet, precluded use of the standard shroud on the Emscope specimen transfer device. Consequently, the sample became heavily contaminated (Fig. 1). A removable shroud was devised and used to reduce contamination (Fig. 2), but the specimen lacked clean fractured edges. This result suggested that low vacuum sputter coating was also limiting resolution.

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