A systems biology approach identifies a regulatory network in parotid acinar cell terminal differentiation.
OBJECTIVE
The transcription factor networks that drive parotid salivary gland progenitor cells to terminally differentiate, remain largely unknown and are vital to understanding the regeneration process.


METHODS
A systems biology approach was taken to measure mRNA and microRNA expression in vivo across acinar cell terminal differentiation in the rat parotid salivary gland. Laser capture microdissection (LCM) was used to specifically isolate acinar cell RNA at times spanning the month-long period of parotid differentiation.


RESULTS
Clustering of microarray measurements suggests that expression occurs in four stages. mRNA expression patterns suggest a novel role for Pparg which is transiently increased during mid postnatal differentiation in concert with several target gene mRNAs. 79 microRNAs are significantly differentially expressed across time. Profiles of statistically significant changes of mRNA expression, combined with reciprocal correlations of microRNAs and their target mRNAs, suggest a putative network involving Klf4, a differentiation inhibiting transcription factor, which decreases as several targeting microRNAs increase late in differentiation. The network suggests a molecular switch (involving Prdm1, Sox11, Pax5, miR-200a, and miR-30a) progressively decreases repression of Xbp1 gene transcription, in concert with decreased translational repression by miR-214. The transcription factor Xbp1 mRNA is initially low, increases progressively, and may be maintained by a positive feedback loop with Atf6. Transfection studies show that Xbp1 activates the Mist1 promoter [corrected]. In addition, Xbp1 and Mist1 each activate the parotid secretory protein (Psp) gene, which encodes an abundant salivary protein, and is a marker of terminal differentiation.


CONCLUSIONS
This study identifies novel expression patterns of Pparg, Klf4, and Sox11 during parotid acinar cell differentiation, as well as numerous differentially expressed microRNAs. Network analysis identifies a novel stemness arm, a genetic switch involving transcription factors and microRNAs, and transition to an Xbp1 driven differentiation network. This proposed network suggests key regulatory interactions in parotid gland terminal differentiation.
Department of Oral Immunology and Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America; Department of Biochemistry & Molecular Biology, University of Louisville, Louisville, Kentucky, United States of America; Institute for Molecular Diversity and Drug Design, University of Louisville, Louisville, Kentucky, United States of America and.
Melissa A
MA
Metzler
Department of Oral Immunology and Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America; Institute for Molecular Diversity and Drug Design, University of Louisville, Louisville, Kentucky, United States of America and.
Srirangapatnam G
SG
Venkatesh
Department of Oral Immunology and Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America.
Jaganathan
J
Lakshmanan
Department of Oral Immunology and Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America.
Anne L
AL
Carenbauer
Department of Oral Immunology and Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America.
Sara M
SM
Perez
Department of Biochemistry & Molecular Biology, University of Louisville, Louisville, Kentucky, United States of America; Institute for Molecular Diversity and Drug Design, University of Louisville, Louisville, Kentucky, United States of America and.
Sarah A
SA
Andres
Department of Bioinformatics and Biostatistics, University of Louisville, Louisville, Kentucky, United States of America.
Savitri
S
Appana
Department of Bioinformatics and Biostatistics, University of Louisville, Louisville, Kentucky, United States of America.
Guy N
GN
Brock
Department of Biochemistry & Molecular Biology, University of Louisville, Louisville, Kentucky, United States of America; Institute for Molecular Diversity and Drug Design, University of Louisville, Louisville, Kentucky, United States of America and.
James L
JL
Wittliff
Department of Oral Immunology and Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America; Department of Biochemistry & Molecular Biology, University of Louisville, Louisville, Kentucky, United States of America; Institute for Molecular Diversity and Drug Design, University of Louisville, Louisville, Kentucky, United States of America and.
Douglas S
DS
Darling
D061354:Acinar Cells / Q000166:cytology*; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology; D004268:DNA-Binding Proteins / Q000235:genetics; D005260:Female; D053263:Gene Regulatory Networks / Q000235:genetics; D000090062:Kruppel-Like Factor 4; D010306:Parotid Gland / Q000166:cytology*; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D000071701:Regulatory Factor X Transcription Factors; D049490:Systems Biology / Q000379:methods*; D014157:Transcription Factors / Q000235:genetics; D000071717:X-Box Binding Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D004268:DNA-Binding Proteins; D000090062:Kruppel-Like Factor 4; D012333:RNA, Messenger; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C105074:Bhlha15 protein, rat; C000715469:Klf4 protein, rat; C000605545:Xbp1 protein, rat