17 research outputs found
Shotgun Label-Free Quantitative Proteomics of Water-Deficit-Stressed Midmature Peanut (<i>Arachis hypogaea</i> L.) Seed
Legume seeds and peanuts, in particular,
are an inexpensive source
of plant proteins and edible oil. A comprehensive understanding of
seed metabolism and the effects of water-deficit stress on the incorporation
of the main storage reserves in seeds, such as proteins, fatty acids,
starch, and secondary metabolites, will enhance our ability to improve
seed quality and yield through molecular breeding programs. In the
present study, we employed a label-free quantitative proteomics approach
to study the functional proteins altered in the midmature (65–70
days postanthesis) peanut seed grown under water-deficit stress conditions.
We created a pod-specific proteome database and identified 93 nonredundant,
statistically significant, and differentially expressed proteins between
well-watered and drought-stressed seeds. Mapping of these differential
proteins revealed three candidate biological pathways (glycolysis,
sucrose and starch metabolism, and fatty acid metabolism) that were
significantly altered due to water-deficit stress. Differential accumulation
of proteins from these pathways provides insight into the molecular
mechanisms underlying the observed physiological changes, which include
reductions in pod yield and biomass, reduced germination, reduced
vigor, decreased seed membrane integrity, increase in storage proteins,
and decreased total fatty acid content. Some of the proteins encoding
rate limiting enzymes of biosynthetic pathways could be utilized by
breeders to improve peanut seed production during water-deficit conditions
in the field. The data have been deposited to the ProteomeXchange
with identifier PXD000308
Shotgun Label-Free Quantitative Proteomics of Water-Deficit-Stressed Midmature Peanut (<i>Arachis hypogaea</i> L.) Seed
Legume seeds and peanuts, in particular,
are an inexpensive source
of plant proteins and edible oil. A comprehensive understanding of
seed metabolism and the effects of water-deficit stress on the incorporation
of the main storage reserves in seeds, such as proteins, fatty acids,
starch, and secondary metabolites, will enhance our ability to improve
seed quality and yield through molecular breeding programs. In the
present study, we employed a label-free quantitative proteomics approach
to study the functional proteins altered in the midmature (65–70
days postanthesis) peanut seed grown under water-deficit stress conditions.
We created a pod-specific proteome database and identified 93 nonredundant,
statistically significant, and differentially expressed proteins between
well-watered and drought-stressed seeds. Mapping of these differential
proteins revealed three candidate biological pathways (glycolysis,
sucrose and starch metabolism, and fatty acid metabolism) that were
significantly altered due to water-deficit stress. Differential accumulation
of proteins from these pathways provides insight into the molecular
mechanisms underlying the observed physiological changes, which include
reductions in pod yield and biomass, reduced germination, reduced
vigor, decreased seed membrane integrity, increase in storage proteins,
and decreased total fatty acid content. Some of the proteins encoding
rate limiting enzymes of biosynthetic pathways could be utilized by
breeders to improve peanut seed production during water-deficit conditions
in the field. The data have been deposited to the ProteomeXchange
with identifier PXD000308
Low Focal Adhesion Signaling Promotes Ground State Pluripotency of Mouse Embryonic Stem Cells
Mouse embryonic stem cells (mESCs)
can be maintained in a pluripotent
state when cultured with 2 inhibitors (2i) of extracellular signal-regulated
kinase (MEK) and glycogen synthase kinase-3 (GSK3), and Royan 2 inhibitors
(R2i) of FGF4 and TGFβ. The molecular mechanisms that control
ESC self-renewal and pluripotency are more important for translating
stem cell technologies to clinical applications. In this study, we
used the shotgun proteomics technique to compare the proteome of the
ground state condition (R2i- and 2i-grown cells) to that of serum.
Out of 1749 proteins identified, 171 proteins were differentially
expressed (<i>p</i> < 0.05) in the 2i, R2i, and serum
samples. Gene ontology (GO) analysis of differentially abundant proteins
showed that the focal adhesion signaling pathway significantly down-regulated
under ground state conditions. mESCs had highly adhesive attachment
under the serum condition, whereas in the 2i and R2i culture conditions,
a loss of adhesion was observed and the cells were rounded and grew
in compact colonies on gelatin. Quantitative RT-PCR showed reduced
expression of the integrins family in the 2i and R2i conditions. The
serum culture had more prominent phosphorylation of focal adhesion
kinase (FAK) compared to 2i and R2i cultures. Activity of the extracellular
signal-regulated kinase (ERK)Â1/2 decreased in the 2i and R2i cultures
compared to serum. Activation of integrins by Mn<sup>2+</sup> in the
2i and R2i cultures resulted in reduced <i>Nanog</i> and
increased the expression of lineage marker genes. In this study, we
demonstrated that reduced focal adhesion enabled mESCs to be maintained
in an undifferentiated and pluripotent state
<i>DDX3Y</i>, a Male-Specific Region of Y Chromosome Gene, May Modulate Neuronal Differentiation
Although
it is apparent that chromosome complement mediates sexually
dimorphic expression patterns of some proteins that lead to functional
differences, there has been insufficient evidence following the manipulation
of the male-specific region of the Y chromosome (MSY) gene expression
during neural development. In this study, we profiled the expression
of 23 MSY genes and 15 of their X-linked homologues during neural
cell differentiation of NTERA-2 human embryonal carcinoma cell line
(NT2) cells in three different developmental stages using qRT-PCR,
Western blotting, and immunofluorescence. The expression level of
12 Y-linked genes significantly increased over neural differentiation,
including <i>RBMY1</i>, <i>EIF1AY</i>, <i>DDX3Y</i>, <i>HSFY1</i>, <i>BPY2</i>,<i> PCDH11Y</i>, <i>UTY</i>, <i>RPS4Y1</i>, <i>USP9Y</i>, <i>SRY</i>, <i>PRY</i>, and <i>ZFY</i>. We showed that siRNA-mediated knockdown of DDX3Y, a
DEAD box RNA helicase enzyme, in neural progenitor cells impaired
cell cycle progression and increased apoptosis, consequently interrupting
differentiation. Label-free quantitative shotgun proteomics based
on a spectral counting approach was then used to characterize the
proteomic profile of the cells after <i>DDX3Y</i> knockdown.
Among 917 reproducibly identified proteins detected, 71 proteins were
differentially expressed following <i>DDX3Y</i> siRNA treatment
compared with mock treated cells. Functional grouping indicated that
these proteins were involved in cell cycle, RNA splicing, and apoptosis,
among other biological functions. Our results suggest that MSY genes
may play an important role in neural differentiation and demonstrate
that <i>DDX3Y</i> could play a multifunctional role in neural
cell development, probably in a sexually dimorphic manner
Shotgun Proteomic Analysis of Long-distance Drought Signaling in Rice Roots
Rice (<i>Oryza sativa</i> L. cv. IR64) was grown in split-root systems to analyze long-distance drought signaling within root systems. This in turn underpins how root systems in heterogeneous soils adapt to drought. The approach was to compare four root tissues: (1) fully watered; (2) fully droughted and split-root systems where (3) one-half was watered and (4) the other half was droughted. This was specifically aimed at identifying how droughted root tissues altered the proteome of adjacent wet roots by hormone signals and how wet roots reciprocally affected dry roots hydraulically. Quantitative label-free shotgun proteomic analysis of four different root tissues resulted in identification of 1487 nonredundant proteins, with nearly 900 proteins present in triplicate in each treatment. Drought caused surprising changes in expression, most notably in partially droughted roots where 38% of proteins were altered in level compared to adjacent watered roots. Specific functional groups changed consistently in drought. Pathogenesis-related proteins were generally up-regulated in response to drought and heat-shock proteins were totally absent in roots of fully watered plants. Proteins involved in transport and oxidation–reduction reactions were also highly dependent upon drought signals, with the former largely absent in roots receiving a drought signal while oxidation–reduction proteins were strongly present during drought. Finally, two functionally contrasting protein families were compared to validate our approach, showing that nine tubulins were strongly reduced in droughted roots while six chitinases were up-regulated, even when the signal arrived remotely from adjacent droughted roots
Shotgun Proteomic Analysis of Long-distance Drought Signaling in Rice Roots
Rice (<i>Oryza sativa</i> L. cv. IR64) was grown in split-root systems to analyze long-distance drought signaling within root systems. This in turn underpins how root systems in heterogeneous soils adapt to drought. The approach was to compare four root tissues: (1) fully watered; (2) fully droughted and split-root systems where (3) one-half was watered and (4) the other half was droughted. This was specifically aimed at identifying how droughted root tissues altered the proteome of adjacent wet roots by hormone signals and how wet roots reciprocally affected dry roots hydraulically. Quantitative label-free shotgun proteomic analysis of four different root tissues resulted in identification of 1487 nonredundant proteins, with nearly 900 proteins present in triplicate in each treatment. Drought caused surprising changes in expression, most notably in partially droughted roots where 38% of proteins were altered in level compared to adjacent watered roots. Specific functional groups changed consistently in drought. Pathogenesis-related proteins were generally up-regulated in response to drought and heat-shock proteins were totally absent in roots of fully watered plants. Proteins involved in transport and oxidation–reduction reactions were also highly dependent upon drought signals, with the former largely absent in roots receiving a drought signal while oxidation–reduction proteins were strongly present during drought. Finally, two functionally contrasting protein families were compared to validate our approach, showing that nine tubulins were strongly reduced in droughted roots while six chitinases were up-regulated, even when the signal arrived remotely from adjacent droughted roots
Shotgun Proteomic Analysis of the Mexican Lime Tree Infected with “<i>Candidatus</i> <i>Phytoplasma aurantifolia</i>”
Infection of Mexican lime trees (<i>Citrus aurantifolia</i> L.) with the specialized bacterium “<i>Candidatus</i> <i>Phytoplasma aurantifolia</i>” causes witches’
broom disease. Witches’ broom disease has the potential to
cause significant economic losses throughout western Asia and North
Africa. We used label-free quantitative shotgun proteomics to study
changes in the proteome of Mexican lime trees in response to infection
by “<i>Ca</i>. <i>Phytoplasma aurantifolia</i>”. Of 990 proteins present in five replicates of healthy and
infected plants, the abundances of 448 proteins changed significantly
in response to phytoplasma infection. Of these, 274 proteins were
less abundant in infected plants than in healthy plants, and 174 proteins
were more abundant in infected plants than in healthy plants. These
448 proteins were involved in stress response, metabolism, growth
and development, signal transduction, photosynthesis, cell cycle,
and cell wall organization. Our results suggest that proteomic changes
in response to infection by phytoplasmas might support phytoplasma
nutrition by promoting alterations in the host’s sugar metabolism,
cell wall biosynthesis, and expression of defense-related proteins.
Regulation of defense-related pathways suggests that defense compounds
are induced in interactions with susceptible as well as resistant
hosts, with the main differences between the two interactions being
the speed and intensity of the response
Shotgun Proteomic Analysis of the Mexican Lime Tree Infected with “<i>Candidatus</i> <i>Phytoplasma aurantifolia</i>”
Infection of Mexican lime trees (<i>Citrus aurantifolia</i> L.) with the specialized bacterium “<i>Candidatus</i> <i>Phytoplasma aurantifolia</i>” causes witches’
broom disease. Witches’ broom disease has the potential to
cause significant economic losses throughout western Asia and North
Africa. We used label-free quantitative shotgun proteomics to study
changes in the proteome of Mexican lime trees in response to infection
by “<i>Ca</i>. <i>Phytoplasma aurantifolia</i>”. Of 990 proteins present in five replicates of healthy and
infected plants, the abundances of 448 proteins changed significantly
in response to phytoplasma infection. Of these, 274 proteins were
less abundant in infected plants than in healthy plants, and 174 proteins
were more abundant in infected plants than in healthy plants. These
448 proteins were involved in stress response, metabolism, growth
and development, signal transduction, photosynthesis, cell cycle,
and cell wall organization. Our results suggest that proteomic changes
in response to infection by phytoplasmas might support phytoplasma
nutrition by promoting alterations in the host’s sugar metabolism,
cell wall biosynthesis, and expression of defense-related proteins.
Regulation of defense-related pathways suggests that defense compounds
are induced in interactions with susceptible as well as resistant
hosts, with the main differences between the two interactions being
the speed and intensity of the response
<i>DDX3Y</i>, a Male-Specific Region of Y Chromosome Gene, May Modulate Neuronal Differentiation
Although
it is apparent that chromosome complement mediates sexually
dimorphic expression patterns of some proteins that lead to functional
differences, there has been insufficient evidence following the manipulation
of the male-specific region of the Y chromosome (MSY) gene expression
during neural development. In this study, we profiled the expression
of 23 MSY genes and 15 of their X-linked homologues during neural
cell differentiation of NTERA-2 human embryonal carcinoma cell line
(NT2) cells in three different developmental stages using qRT-PCR,
Western blotting, and immunofluorescence. The expression level of
12 Y-linked genes significantly increased over neural differentiation,
including <i>RBMY1</i>, <i>EIF1AY</i>, <i>DDX3Y</i>, <i>HSFY1</i>, <i>BPY2</i>,<i> PCDH11Y</i>, <i>UTY</i>, <i>RPS4Y1</i>, <i>USP9Y</i>, <i>SRY</i>, <i>PRY</i>, and <i>ZFY</i>. We showed that siRNA-mediated knockdown of DDX3Y, a
DEAD box RNA helicase enzyme, in neural progenitor cells impaired
cell cycle progression and increased apoptosis, consequently interrupting
differentiation. Label-free quantitative shotgun proteomics based
on a spectral counting approach was then used to characterize the
proteomic profile of the cells after <i>DDX3Y</i> knockdown.
Among 917 reproducibly identified proteins detected, 71 proteins were
differentially expressed following <i>DDX3Y</i> siRNA treatment
compared with mock treated cells. Functional grouping indicated that
these proteins were involved in cell cycle, RNA splicing, and apoptosis,
among other biological functions. Our results suggest that MSY genes
may play an important role in neural differentiation and demonstrate
that <i>DDX3Y</i> could play a multifunctional role in neural
cell development, probably in a sexually dimorphic manner
Shotgun Proteomic Analysis of the Mexican Lime Tree Infected with “<i>Candidatus</i> <i>Phytoplasma aurantifolia</i>”
Infection of Mexican lime trees (<i>Citrus aurantifolia</i> L.) with the specialized bacterium “<i>Candidatus</i> <i>Phytoplasma aurantifolia</i>” causes witches’
broom disease. Witches’ broom disease has the potential to
cause significant economic losses throughout western Asia and North
Africa. We used label-free quantitative shotgun proteomics to study
changes in the proteome of Mexican lime trees in response to infection
by “<i>Ca</i>. <i>Phytoplasma aurantifolia</i>”. Of 990 proteins present in five replicates of healthy and
infected plants, the abundances of 448 proteins changed significantly
in response to phytoplasma infection. Of these, 274 proteins were
less abundant in infected plants than in healthy plants, and 174 proteins
were more abundant in infected plants than in healthy plants. These
448 proteins were involved in stress response, metabolism, growth
and development, signal transduction, photosynthesis, cell cycle,
and cell wall organization. Our results suggest that proteomic changes
in response to infection by phytoplasmas might support phytoplasma
nutrition by promoting alterations in the host’s sugar metabolism,
cell wall biosynthesis, and expression of defense-related proteins.
Regulation of defense-related pathways suggests that defense compounds
are induced in interactions with susceptible as well as resistant
hosts, with the main differences between the two interactions being
the speed and intensity of the response